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Kara MF, Guo W, Zhang R, Denby K. LsRTDv1, a reference transcript dataset for accurate transcript-specific expression analysis in lettuce. THE PLANT JOURNAL : FOR CELL AND MOLECULAR BIOLOGY 2024; 120:370-386. [PMID: 39145419 DOI: 10.1111/tpj.16978] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/25/2024] [Revised: 06/20/2024] [Accepted: 07/31/2024] [Indexed: 08/16/2024]
Abstract
Accurate quantification of gene and transcript-specific expression, with the underlying knowledge of precise transcript isoforms, is crucial to understanding many biological processes. Analysis of RNA sequencing data has benefited from the development of alignment-free algorithms which enhance the precision and speed of expression analysis. However, such algorithms require a reference transcriptome. Here we generate a reference transcript dataset (LsRTDv1) for lettuce (cv. Saladin), combining long- and short-read sequencing with publicly available transcriptome annotations, and filtering to keep only transcripts with high-confidence splice junctions and transcriptional start and end sites. LsRTDv1 identifies novel genes (mostly long non-coding RNAs) and increases the number of transcript isoforms per gene in the lettuce genome from 1.4 to 2.7. We show that LsRTDv1 significantly increases the mapping rate of RNA-seq data from a lettuce time-series experiment (mock- and Botrytis cinerea-inoculated) and enables detection of genes that are differentially alternatively spliced in response to infection as well as transcript-specific expression changes. LsRTDv1 is a valuable resource for investigation of transcriptional and alternative splicing regulation in lettuce.
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Affiliation(s)
- Mehmet Fatih Kara
- Biology Department, Centre for Novel Agricultural Products (CNAP), University of York, Wentworth Way, York, YO10 5DD, UK
| | - Wenbin Guo
- Information and Computational Sciences, James Hutton Institute, Dundee, DD2 5DA, UK
| | - Runxuan Zhang
- Information and Computational Sciences, James Hutton Institute, Dundee, DD2 5DA, UK
| | - Katherine Denby
- Biology Department, Centre for Novel Agricultural Products (CNAP), University of York, Wentworth Way, York, YO10 5DD, UK
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2
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Liu X, Li M, Chen T, Zhang R, Wang Y, Xiao J, Ding X, Zhang S, Li Q. A global survey of bicarbonate stress-induced pre-mRNA alternative splicing in soybean via integrative analysis of Iso-seq and RNA-seq. Int J Biol Macromol 2024; 278:135067. [PMID: 39191343 DOI: 10.1016/j.ijbiomac.2024.135067] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2024] [Revised: 08/23/2024] [Accepted: 08/23/2024] [Indexed: 08/29/2024]
Abstract
Alternative splicing (AS) plays important roles in modulating environmental stress responses in plants. However, little is known about the functions of bicarbonate-induced AS in cultivated soybean (Glycine max L. Merr.). In this study, we combined PacBio isoform sequencing (Iso-seq) and Illumina RNA sequencing (RNA-seq) to elucidate the bicarbonate-induced AS events in soybean root and leaf tissues. Compared to RNA-seq, Iso-seq identified more novel genes and transcripts, as well as more AS events, indicating that Iso-seq is more efficient in AS detection. Combining these two technologies, we found that intron retention (IR) is the most frequent AS event type. We identified a total of 913 and 1974 bicarbonate stress-responsive differentially alternative spliced genes (DAGs) in soybean leaves and roots respectively, from our RNA-seq results. Additionally, we determined a transcription factor (GmNTL9) and a splicing factor (GmRSZ22), and validated their roles in bicarbonate stress response by AS. Overall, our study opens an avenue for evaluating plant AS regulatory networks, and the obtained global landscape of alternative splicing provides valuable insights into the AS-mediated bicarbonate-responsive mechanisms in plant species.
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Affiliation(s)
- Xin Liu
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China; Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin 150030, China
| | - Minglong Li
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China; Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin 150030, China
| | - Tong Chen
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China
| | - Rui Zhang
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China
| | - Yuye Wang
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China
| | - Jialei Xiao
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China; Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin 150030, China
| | - Xiaodong Ding
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China; Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin 150030, China.
| | - Shuzhen Zhang
- Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin 150030, China; Department of Plant Science, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai 200240, China.
| | - Qiang Li
- Key Laboratory of Agricultural Biological Functional Genes, Northeast Agricultural University, Harbin 150030, China; Key Laboratory of Soybean Biology of Chinese Education Ministry, Northeast Agricultural University, Harbin 150030, China.
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Wang X, Wang X, Mu H, Zhao B, Song X, Fan H, Wang B, Yuan F. Global analysis of key post-transcriptional regulation in early leaf development of Limonium bicolor identifies a long non-coding RNA that promotes salt gland development and salt resistance. JOURNAL OF EXPERIMENTAL BOTANY 2024; 75:5091-5110. [PMID: 38795330 DOI: 10.1093/jxb/erae241] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/15/2023] [Accepted: 05/23/2024] [Indexed: 05/27/2024]
Abstract
Limonium bicolor, known horticulturally as sea lavender, is a typical recretohalophyte with salt glands in its leaf epidermis that secrete excess Na+ out of the plant. Although many genes have been proposed to contribute to salt gland initiation and development, a detailed analysis of alternative splicing, alternative polyadenylation patterns, and long non-coding RNAs (lncRNAs) has been lacking. Here, we applied single-molecule long-read mRNA isoform sequencing (Iso-seq) to explore the complexity of the L. bicolor transcriptome in leaves during salt gland initiation (stage A) and salt gland differentiation (stage B) based on the reference genome. We identified alternative splicing events and the use of alternative poly(A) sites unique to stage A or stage B, leading to the hypothesis that they might contribute to the differentiation of salt glands. Based on the Iso-seq data and RNA in situ hybridization of candidate genes, we selected the lncRNA Btranscript_153392 for gene editing and virus-induced gene silencing to dissect its function. In the absence of this transcript, we observed fewer salt glands on the leaf epidermis, leading to diminished salt secretion and salt tolerance. Our data provide transcriptome resources for unraveling the mechanisms behind salt gland development and furthering crop transformation efforts towards enhanced survivability in saline soils.
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Affiliation(s)
- Xi Wang
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Xiaoyu Wang
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Huiying Mu
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Boqing Zhao
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Xianrui Song
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Hai Fan
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Baoshan Wang
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
| | - Fang Yuan
- Shandong Provincial Key Laboratory of Plant Stress, College of Life Sciences, Shandong Normal University, Ji'nan, Shandong, China
- National Center of Technology Innovation for Comprehensive Utilization of Saline-Alkali Land, Dongying, Shandong, China
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4
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Shi ZX, Xiang L, Zhao HM, Yang LQ, Chen ZC, Pu YQ, Li YW, Luo B, Cai QY, Liu BL, Feng NX, Li H, Li QX, Tang C, Mo CH. High-throughput single-molecule long-read RNA sequencing analysis of tissue-specific genes and isoforms in lettuce (Lactuca sativa L.). Commun Biol 2024; 7:920. [PMID: 39080448 PMCID: PMC11289301 DOI: 10.1038/s42003-024-06598-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/17/2024] [Accepted: 07/17/2024] [Indexed: 08/02/2024] Open
Abstract
Lettuce is one of the most widely cultivated and consumed dicotyledonous vegetables globally. Despite the availability of its reference genome sequence, lettuce gene annotation remains incomplete, impeding comprehensive research and the broad application of genomic resources. Long-read RNA isoform sequencing (Iso-Seq) offers substantial advantages for analyzing RNA alternative splicing and aiding gene annotation, yet it faces throughput limitations. We present the HIT-ISOseq method tailored for bulk sample analysis, significantly enhancing RNA sequencing throughput on the PacBio platform by concatenating cDNA. Here we show, HIT-ISOseq generates 3-4 cDNA molecules per CCS read in lettuce, yielding 15.7 million long reads per PacBio Sequel II SMRT Cell 8 M. We validate its effectiveness in analyzing six lettuce tissue samples, including roots, stems, and leaves, revealing tissue-specific gene expression patterns and RNA isoforms. Leveraging diverse tissue long-read RNA sequencing, we refine the transcript annotation of the lettuce reference genome, expanding its GO and KEGG annotation repertoire. Collectively, this study serves as a foundational reference for genome annotation and the analysis of multi-sample isoform expression, utilizing high-throughput long-read transcriptome sequencing.
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Affiliation(s)
- Zhuo-Xing Shi
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
- BGI Genomics, BGI-Shenzhen, Shenzhen, China
| | - Lei Xiang
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Hai-Ming Zhao
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Lang-Qi Yang
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | | | - Yu-Qing Pu
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Yan-Wen Li
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Bei Luo
- BGI Genomics, BGI-Shenzhen, Shenzhen, China
| | - Quan-Ying Cai
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Bai-Lin Liu
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Nai-Xian Feng
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Hui Li
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China
| | - Qing X Li
- Department of Molecular Biosciences and Bioengineering, University of Hawaii at Manoa, Honolulu, HI, 96822, USA
| | - Chong Tang
- BGI Genomics, BGI-Shenzhen, Shenzhen, China.
| | - Ce-Hui Mo
- Guangdong Provincial Research Center for Environment Pollution Control and Remediation Materials, College of Life Science and Technology, Jinan University, Guangzhou, 510632, China.
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Dong X, Li W, Li C, Akan OD, Liao C, Cao J, Zhang L. Integrated transcriptomics and metabolomics revealed the mechanism of catechin biosynthesis in response to lead stress in tung tree (Vernicia fordii). THE SCIENCE OF THE TOTAL ENVIRONMENT 2024; 930:172796. [PMID: 38692325 DOI: 10.1016/j.scitotenv.2024.172796] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/15/2024] [Revised: 04/24/2024] [Accepted: 04/24/2024] [Indexed: 05/03/2024]
Abstract
Lead (Pb) affects gene transcription, metabolite biosynthesis and growth in plants. The tung tree (Vernicia fordii) is highly adaptive to adversity, whereas the mechanisms underlying its response to Pb remain uncertain. In this work, transcriptomic and metabolomic analyses were employed to study tung trees under Pb stress. The results showed that the biomass of tung seedlings decreased with increasing Pb doses, and excessive Pb doses resulted in leaf wilting, root rot, and disruption of Pb homeostasis. Under non-excessive Pb stress, a significant change in the expression patterns of flavonoid biosynthesis genes was observed in the roots of tung seedlings, leading to changes in the accumulation of flavonoids in the roots, especially the upregulation of catechins, which can chelate Pb and reduce its toxicity in plants. In addition, Pb-stressed roots showed a large accumulation of VfWRKY55, VfWRKY75, and VfLRR1 transcripts, which were shown to be involved in the flavonoid biosynthesis pathway by gene module analysis. Overexpression of VfWRKY55, VfWRKY75, and VfLRR1 significantly increased catechin concentrations in tung roots, respectively. These data indicate that Pb stress-induced changes in the expression patterns of those genes regulate the accumulation of catechins. Our findings will help to clarify the molecular mechanism of Pb response in plants.
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Affiliation(s)
- Xiang Dong
- Key Laboratory of Cultivation and Protection for Non-wood Forest Trees, Ministry of Education, Central South University of Forestry and Technology, Changsha 410004, China
| | - Wenying Li
- Key Laboratory of Cultivation and Protection for Non-wood Forest Trees, Ministry of Education, Central South University of Forestry and Technology, Changsha 410004, China; College of Biology and Agricultural Resources, Huanggang Normal University, Huanggang, Hubei 438000, China
| | - Changzhu Li
- State Key Laboratory of Utilization of Woody Oil Resource, Hunan Academy of Forestry, Changsha 410004, China
| | - Otobong Donald Akan
- Key Laboratory of Cultivation and Protection for Non-wood Forest Trees, Ministry of Education, Central South University of Forestry and Technology, Changsha 410004, China; Faculty of Biological Science, Akwa-Ibom State University, Akwa-Ibom State, Uyo 1167, Nigeria
| | - Chancan Liao
- Key Laboratory of Cultivation and Protection for Non-wood Forest Trees, Ministry of Education, Central South University of Forestry and Technology, Changsha 410004, China
| | - Jie Cao
- Key Laboratory of Cultivation and Protection for Non-wood Forest Trees, Ministry of Education, Central South University of Forestry and Technology, Changsha 410004, China
| | - Lin Zhang
- Key Laboratory of Cultivation and Protection for Non-wood Forest Trees, Ministry of Education, Central South University of Forestry and Technology, Changsha 410004, China.
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Greshnova A, Pál K, Martinez JFI, Canzar S, Makova KD. Transcript Isoform Diversity of Y Chromosome Ampliconic Genes of Great Apes Uncovered Using Long Reads and Telomere-to-Telomere Reference Genome Assemblies. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.04.02.587783. [PMID: 38617276 PMCID: PMC11014635 DOI: 10.1101/2024.04.02.587783] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/16/2024]
Abstract
Y chromosomes of great apes harbor Ampliconic Genes (YAGs)-multi-copy gene families (BPY2, CDY, DAZ, HSFY, PRY, RBMY, TSPY, VCY, and XKRY) that encode proteins important for spermatogenesis. Previous work assembled YAG transcripts based on their targeted sequencing but not using reference genome assemblies, potentially resulting in an incomplete transcript repertoire. Here we used the recently produced gapless telomere-to-telomere (T2T) Y chromosome assemblies of great ape species (bonobo, chimpanzee, human, gorilla, Bornean orangutan, and Sumatran orangutan) and analyzed RNA data from whole-testis samples for the same species. We generated hybrid transcriptome assemblies by combining targeted long reads (Pacific Biosciences), untargeted long reads (Pacific Biosciences) and untargeted short reads (Illumina)and mapping them to the T2T reference genomes. Compared to the results from the reference-free approach, average transcript length was more than two times higher, and the total number of transcripts decreased three times, improving the quality of the assembled transcriptome. The reference-based transcriptome assemblies allowed us to differentiate transcripts originating from different Y chromosome gene copies and from their non-Y chromosome homologs. We identified two sources of transcriptome diversity-alternative splicing and gene duplication with subsequent diversification of gene copies. For each gene family, we detected transcribed pseudogenes along with protein-coding gene copies. We revealed previously unannotated gene copies of YAGs as compared to currently available NCBI annotations, as well as novel isoforms for annotated gene copies. This analysis paves the way for better understanding Y chromosome gene functions, which is important given their role in spermatogenesis.
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Affiliation(s)
- Aleksandra Greshnova
- Department of Biology, Penn State University, University Park, PA, USA
- Current address: Max Planck Institute for Evolutionary Biology, Plön, Germany
| | - Karol Pál
- Department of Biology, Penn State University, University Park, PA, USA
| | - Juan Francisco Iturralde Martinez
- Department of Computer Science and Engineering, The Pennsylvania State University, University Park, PA 16802, United States
- Huck Institutes of the Life Sciences. Pennsylvania State University, University Park, PA 16802, USA
| | - Stefan Canzar
- Faculty of Informatics and Data Science, University of Regensburg, Regensburg, Germany
- Department of Computer Science and Engineering, The Pennsylvania State University, University Park, PA 16802, United States
| | - Kateryna D Makova
- Department of Biology, Penn State University, University Park, PA, USA
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Sena L, Mica E, Valè G, Vaccino P, Pecchioni N. Exploring the potential of endophyte-plant interactions for improving crop sustainable yields in a changing climate. FRONTIERS IN PLANT SCIENCE 2024; 15:1349401. [PMID: 38571718 PMCID: PMC10988515 DOI: 10.3389/fpls.2024.1349401] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 12/04/2023] [Accepted: 03/05/2024] [Indexed: 04/05/2024]
Abstract
Climate change poses a major threat to global food security, significantly reducing crop yields as cause of abiotic stresses, and for boosting the spread of new and old pathogens and pests. Sustainable crop management as a route to mitigation poses the challenge of recruiting an array of solutions and tools for the new aims. Among these, the deployment of positive interactions between the micro-biotic components of agroecosystems and plants can play a highly significant role, as part of the agro-ecological revolution. Endophytic microorganisms have emerged as a promising solution to tackle this challenge. Among these, Arbuscular Mycorrhizal Fungi (AMF) and endophytic bacteria and fungi have demonstrated their potential to alleviate abiotic stresses such as drought and heat stress, as well as the impacts of biotic stresses. They can enhance crop yields in a sustainable way also by other mechanisms, such as improving the nutrient uptake, or by direct effects on plant physiology. In this review we summarize and update on the main types of endophytes, we highlight several studies that demonstrate their efficacy in improving sustainable yields and explore possible avenues for implementing crop-microbiota interactions. The mechanisms underlying these interactions are highly complex and require a comprehensive understanding. For this reason, omic technologies such as genomics, transcriptomics, proteomics, and metabolomics have been employed to unravel, by a higher level of information, the complex network of interactions between plants and microorganisms. Therefore, we also discuss the various omic approaches and techniques that have been used so far to study plant-endophyte interactions.
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Affiliation(s)
- Lorenzo Sena
- Dipartimento di Scienze della Vita, Sede Agraria, UNIMORE - Università di Modena e Reggio Emilia, Reggio Emilia, Italy
- Centro di Ricerca Cerealicoltura e Colture Industriali, CREA – Consiglio per la Ricerca in Agricoltura e l’Analisi dell’Economia Agraria, Vercelli, Italy
| | - Erica Mica
- Dipartimento per lo Sviluppo Sostenibile e la Transizione Ecologica, UPO – Università del Piemonte Orientale, Complesso San Giuseppe, Vercelli, Italy
| | - Giampiero Valè
- Dipartimento per lo Sviluppo Sostenibile e la Transizione Ecologica, UPO – Università del Piemonte Orientale, Complesso San Giuseppe, Vercelli, Italy
| | - Patrizia Vaccino
- Centro di Ricerca Cerealicoltura e Colture Industriali, CREA – Consiglio per la Ricerca in Agricoltura e l’Analisi dell’Economia Agraria, Vercelli, Italy
| | - Nicola Pecchioni
- Dipartimento di Scienze della Vita, Sede Agraria, UNIMORE - Università di Modena e Reggio Emilia, Reggio Emilia, Italy
- Centro di Ricerca Cerealicoltura e Colture Industriali, CREA – Consiglio per la Ricerca in Agricoltura e l’Analisi dell’Economia Agraria, Vercelli, Italy
- Centro di Ricerca Cerealicoltura e Colture Industriali, CREA – Consiglio per la Ricerca in Agricoltura e l’Analisi dell’Economia Agraria, Foggia, Italy
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Kang JN, Hur M, Kim CK, Yang SH, Lee SM. Enhancing transcriptome analysis in medicinal plants: multiple unigene sets in Astragalus membranaceus. FRONTIERS IN PLANT SCIENCE 2024; 15:1301526. [PMID: 38384760 PMCID: PMC10879423 DOI: 10.3389/fpls.2024.1301526] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 09/26/2023] [Accepted: 01/22/2024] [Indexed: 02/23/2024]
Abstract
Astragalus membranaceus is a medicinal plant mainly used in East Asia and contains abundant secondary metabolites. Despite the importance of this plant, the available genomic and genetic information is still limited. De novo transcriptome construction is recognized as an essential method for transcriptome research when reference genome information is incomplete. In this study, we constructed three individual transcriptome sets (unigene sets) for detailed analysis of the phenylpropanoid biosynthesis pathway, a major metabolite of A. membranaceus. Set-1 was a circular consensus sequence (CCS) generated using PacBio sequencing (PacBio-seq). Set-2 consisted of hybridized assembled unigenes with Illumina sequencing (Illumina-seq) reads and PacBio CCS using rnaSPAdes. Set-3 unigenes were assembled from Illumina-seq reads using the Trinity software. Construction of multiple unigene sets provides several advantages for transcriptome analysis. First, it provides an appropriate expression filtering threshold for assembly-based unigenes: a threshold transcripts per million (TPM) ≥ 5 removed more than 88% of assembly-based unigenes, which were mostly short and low-expressing unigenes. Second, assembly-based unigenes compensated for the incomplete length of PacBio CCSs: the ends of the 5`/3` untranslated regions of phenylpropanoid-related unigenes derived from set-1 were incomplete, which suggests that PacBio CCSs are unlikely to be full-length transcripts. Third, more isoform unigenes could be obtained from multiple unigene sets; isoform unigenes missing in Set-1 were detected in set-2 and set-3. Finally, gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses showed that phenylpropanoid biosynthesis and carbohydrate metabolism were highly activated in A. membranaceus roots. Various sequencing technologies and assemblers have been developed for de novo transcriptome analysis. However, no technique is perfect for de novo transcriptome analysis, suggesting the need to construct multiple unigene sets. This method enables efficient transcript filtering and detection of longer and more diverse transcripts.
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Affiliation(s)
- Ji-Nam Kang
- Genomics Division, National Institute of Agricultural Sciences, Jeonju-si, Jeollabuk-do, Republic of Korea
| | - Mok Hur
- Department of Herbal Crop Resources, National Institute of Horticultural & Herbal Science, Eumseong-gun, Chungcheongbuk-do, Republic of Korea
| | - Chang-Kug Kim
- Genomics Division, National Institute of Agricultural Sciences, Jeonju-si, Jeollabuk-do, Republic of Korea
| | - So-Hee Yang
- Genomics Division, National Institute of Agricultural Sciences, Jeonju-si, Jeollabuk-do, Republic of Korea
| | - Si-Myung Lee
- Genomics Division, National Institute of Agricultural Sciences, Jeonju-si, Jeollabuk-do, Republic of Korea
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Zhao X, Yan F, Li YM, Tang J, Hu XC, Feng Z, Gao J, Peng L, Zhang G. Comparative transcriptome analysis and identification of candidate R2R3-MYB genes involved in anthraquinone biosynthesis in Rheum palmatum L. Chin Med 2024; 19:23. [PMID: 38317158 PMCID: PMC10845799 DOI: 10.1186/s13020-024-00891-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2023] [Accepted: 01/21/2024] [Indexed: 02/07/2024] Open
Abstract
BACKGROUND Rheum palmatum L. has important medicinal value because it contains biologically active anthraquinones. However, the key genes and TFs involved in anthraquinone biosynthesis and regulation in R. palmatum remain unclear. METHODS Based on full length transcriptome data, in this study, we screened the differentially expressed genes in the anthraquinone biosynthesis pathway. The R2R3-MYB family genes of R. palmatum were systematically identified based on full-length transcriptome sequencing followed by bioinformatics analyses. The correlation analysis was carried out by using co-expression analysis, protein interaction analysis, and real-time fluorescence quantitative analysis after MeJA treatment. The RpMYB81 and RpMYB98 genes were amplified by RT-PCR, and their subcellular localization and self-activation characteristics were analyzed. RESULTS Comparative transcriptome analysis results revealed a total of 3525 upregulated and 6043 downregulated DEGs in the CK versus MeJA group; 28 DEGs were involved in the anthraquinone pathway. Eleven CHS genes that belonged to the PKS family were differentially expressed and involved in anthraquinone biosynthesis. Twelve differentially expressed MYBs genes were found to be co-expressed and interact with CHS genes. Furthermore, 52 MYB genes were identified as positive regulators of anthraquinone biosynthesis and were further characterized. Three MYB genes including RpMYB81, RpMYB98, and RpMYB100 responded to MeJA treatment in R. palmatum, and the levels of these genes were verified by qRT-PCR. RpMYB81 was related to anthraquinone biosynthesis. RpMYB98 had an interaction with genes in the anthraquinone biosynthesis pathway. RpMYB81 and RpMYB98 were mainly localized in the nucleus. RpMYB81 had self-activation activity, while RpMYB98 had no self-activation activity. CONCLUSION RpMYB81, RpMYB98, and RpMYB100 were significantly induced by MeJA treatment. RpMYB81 and RpMYB98 are located in the nucleus, and RpMYB81 has transcriptional activity, suggesting that it might be involved in the transcriptional regulation of anthraquinone biosynthesis in R. palmatum.
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Affiliation(s)
- Xia Zhao
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Feng Yan
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Yi-Min Li
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China.
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China.
| | - Jing Tang
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Xiao-Chen Hu
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Zhao Feng
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Jing Gao
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Liang Peng
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China
| | - Gang Zhang
- Key Laboratory for Research and Development of "Qin Medicine" of Shaanxi Administration of Traditional Chinese Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China.
- College of Pharmacy and Shaanxi Qinling Application Development and Engineering Center of Chinese Herbal Medicine, Shaanxi University of Chinese Medicine, Xianyang, 712046, China.
- State Key Laboratory of Research and Development of Characteristic Qin Medicine Resources (Cultivation), Shaanxi University of Chinese Medicine, Xianyang, 712083, China.
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Ma Y, Li J, Yu H, Teng L, Geng H, Li R, Xing R, Liu S, Li P. Comparative analysis of PacBio and ONT RNA sequencing methods for Nemopilema Nomurai venom identification. Genomics 2023; 115:110709. [PMID: 37739021 DOI: 10.1016/j.ygeno.2023.110709] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2023] [Revised: 08/28/2023] [Accepted: 09/18/2023] [Indexed: 09/24/2023]
Abstract
Recent studies on marine organisms have made use of third-generation sequencing technologies such as Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT). While these specialized bioinformatics tools have different algorithmic designs and performance capabilities, they offer scalability and can be applied to various datasets. We investigated the effectiveness of PacBio and ONT RNA sequencing methods in identifying the venom of the jellyfish species Nemopilema nomurai. We conducted a detailed analysis of the sequencing data from both methods, focusing on key characteristics such as CD, alternative splicing, long-chain noncoding RNA, simple sequence repeat, transcription factor, and functional transcript annotation. Our findings indicate that ONT generally produced higher raw data quality in the transcriptome analysis, while PacBio generated longer read lengths. PacBio was found to be superior in identifying CDs and long-chain noncoding RNA, whereas ONT was more cost-effective for predicting alternative splicing events, simple sequence repeats, and transcription factors. Based on these results, we conclude that PacBio is the most specific and sensitive method for identifying venom components, while ONT is the most cost-effective method for studying venogenesis, cnidocyst (venom gland) development, and transcription of virulence genes in jellyfish. Our study has implications for future sequencing technologies in marine jellyfish, and highlights the power of full-length transcriptome analysis in discovering potential therapeutic targets for jellyfish dermatitis.
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Affiliation(s)
- Yuzhen Ma
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China
| | - Jie Li
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China; University of Chinese Academy of Sciences, Beijing 100049, China
| | - Huahua Yu
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China.
| | - Lichao Teng
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China; University of Chinese Academy of Sciences, Beijing 100049, China
| | - Hao Geng
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China; University of Chinese Academy of Sciences, Beijing 100049, China
| | - Rongfeng Li
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China
| | - Ronge Xing
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China
| | - Song Liu
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China
| | - Pengcheng Li
- Key Laboratory of Experimental Marine Biology, Center for Ocean Mega-Science, Institute of Oceanology, Chinese Academy of Sciences, Qingdao 266071, China; Laboratory for Marine Drugs and Bioproducts, Pilot National Laboratory for Marine Science and Technology (Qingdao), No. 1 Wenhai Road, Qingdao 266237, China.
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Al-Dossary O, Furtado A, KharabianMasouleh A, Alsubaie B, Al-Mssallem I, Henry RJ. Long read sequencing to reveal the full complexity of a plant transcriptome by targeting both standard and long workflows. PLANT METHODS 2023; 19:112. [PMID: 37865785 PMCID: PMC10589961 DOI: 10.1186/s13007-023-01091-1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/17/2022] [Accepted: 10/13/2023] [Indexed: 10/23/2023]
Abstract
BACKGROUND Long read sequencing allows the analysis of full-length transcripts in plants without the challenges of reliable transcriptome assembly. Long read sequencing of transcripts from plant genomes has often utilized sized transcript libraries. However, the value of including libraries of differing sizes has not been established. METHODS A comprehensive transcriptome of the leaves of Jojoba (Simmondsia chinensis) was generated from two different PacBio library preparations: standard workflow (SW) and long workflow (LW). RESULTS The importance of using both transcript groups in the analysis was demonstrated by the high proportion of unique sequences (74.6%) that were not shared between the groups. A total of 37.8% longer transcripts were only detected in the long dataset. The completeness of the combined transcriptome was indicated by the presence of 98.7% of genes predicted in the jojoba male reference genome. The high coverage of the transcriptome was further confirmed by BUSCO analysis showing the presence of 96.9% of the genes from the core viridiplantae_odb10 lineage. The high-quality isoforms post Cd-Hit merged dataset of the two workflows had a total of 167,866 isoforms. Most of the transcript isoforms were protein-coding sequences (71.7%) containing open reading frames (ORFs) ≥ 100 amino acids (aa). Alternative splicing and intron retention were the basis of most transcript diversity when analysed at the whole genome level and by specific analysis of the apetala2 gene families. CONCLUSION This suggests the need to specifically target the capture of longer transcripts to provide more comprehensive genome coverage in plant transcriptome analysis and reveal the high level of alternative splicing.
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Affiliation(s)
- Othman Al-Dossary
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072, Australia
- College of Agriculture and Food Sciences, King Faisal University, 36362, Al Hofuf, Saudi Arabia
| | - Agnelo Furtado
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072, Australia
| | - Ardashir KharabianMasouleh
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072, Australia
| | - Bader Alsubaie
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072, Australia
- College of Agriculture and Food Sciences, King Faisal University, 36362, Al Hofuf, Saudi Arabia
| | - Ibrahim Al-Mssallem
- College of Agriculture and Food Sciences, King Faisal University, 36362, Al Hofuf, Saudi Arabia
| | - Robert J Henry
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072, Australia.
- ARC Centre of Excellence for Plant Success in Nature and Agriculture, University of Queensland, Brisbane, 4072, Australia.
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12
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Zou Y, Guo Q, Chang Y, Zhong Y, Cheng L, Wei W. Alternative splicing affects synapses in the hippocampus of offspring after maternal fructose exposure during gestation and lactation. Chem Biol Interact 2023; 379:110518. [PMID: 37121297 DOI: 10.1016/j.cbi.2023.110518] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2023] [Revised: 04/15/2023] [Accepted: 04/27/2023] [Indexed: 05/02/2023]
Abstract
Increased fructose over-intake is a global issue. Maternal fructose exposure during gestation and lactation can impair brain development in offspring. However, the effect on synapses is still unknown. For the diversification of RNA and biological functions, alternative splicing (AS) and alternative polyadenylation (APA) are essential. We constructed a maternal high-fructose diet model by administering 13% and 40% fructose water. The student's t-test analyzed the results of RT-qPCR. All other results were analyzed by one-way analysis of variance. The animal behavior experiment results revealed that conditioning and associative memory had been damaged. The proteins that form synapses were consistently low-expressed. In addition, compared with the control group, the Oxford Nanopore Technologies platform's full-length RNA-sequencing identified 298 different spliced genes (DSGs) and 51 differentially expressed alternative splicing (DEAS) genes in the 13% fructose group. 313 DSGs and 74 DEAS genes were in the 40% fructose group. Enrichment analysis based on these altered genes revealed some enlightening items and pathways. Our findings demonstrated the transcriptome mechanism underlying maternal fructose exposure during gestation and lactation and impaired synapse function during the transcripts' editing.
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Affiliation(s)
- Yuchen Zou
- Child and Adolescent Health, School of Public Health, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, PR China
| | - Qing Guo
- Child and Adolescent Health, School of Public Health, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, PR China
| | - Yidan Chang
- Child and Adolescent Health, School of Public Health, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, PR China
| | - Yongyong Zhong
- Child and Adolescent Health, School of Public Health, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, PR China
| | - Lin Cheng
- Child and Adolescent Health, School of Public Health, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, PR China
| | - Wei Wei
- Child and Adolescent Health, School of Public Health, China Medical University, No.77 Puhe Road, Shenyang North New Area, Shenyang, 110122, PR China.
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Sybilska E, Daszkowska-Golec A. Alternative splicing in ABA signaling during seed germination. FRONTIERS IN PLANT SCIENCE 2023; 14:1144990. [PMID: 37008485 PMCID: PMC10060653 DOI: 10.3389/fpls.2023.1144990] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 01/15/2023] [Accepted: 03/02/2023] [Indexed: 06/19/2023]
Abstract
Seed germination is an essential step in a plant's life cycle. It is controlled by complex physiological, biochemical, and molecular mechanisms and external factors. Alternative splicing (AS) is a co-transcriptional mechanism that regulates gene expression and produces multiple mRNA variants from a single gene to modulate transcriptome diversity. However, little is known about the effect of AS on the function of generated protein isoforms. The latest reports indicate that alternative splicing (AS), the relevant mechanism controlling gene expression, plays a significant role in abscisic acid (ABA) signaling. In this review, we present the current state of the art about the identified AS regulators and the ABA-related changes in AS during seed germination. We show how they are connected with the ABA signaling and the seed germination process. We also discuss changes in the structure of the generated AS isoforms and their impact on the functionality of the generated proteins. Also, we point out that the advances in sequencing technology allow for a better explanation of the role of AS in gene regulation by more accurate detection of AS events and identification of full-length splicing isoforms.
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Affiliation(s)
| | - Agata Daszkowska-Golec
- Institute of Biology, Biotechnology and Environmental Protection, Faculty of Natural Sciences, University of Silesia in Katowice, Katowice, Poland
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14
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Li Y, Zhang T, Kang Y, Wang P, Yu W, Wang J, Li W, Jiang X, Zhou Y. Integrated metabolome, transcriptome analysis, and multi-flux full-length sequencing offer novel insights into the function of lignin biosynthesis as a Sesuvium portulacastrum response to salt stress. Int J Biol Macromol 2023; 237:124222. [PMID: 36990407 DOI: 10.1016/j.ijbiomac.2023.124222] [Citation(s) in RCA: 12] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2023] [Revised: 03/21/2023] [Accepted: 03/24/2023] [Indexed: 03/30/2023]
Abstract
Sesuvium portulacastrum is a typical halophyte. However, few studies have investigated its salt-tolerant molecular mechanism. In this study, metabolome, transcriptome, and multi-flux full-length sequencing analysis were conducted to investigate the significantly different metabolites (SDMs) and differentially expressed genes (DEGs) of S. portulacastrum samples under salinity. The complete-length transcriptome of S. portulacastrum was developed, which contained 39,659 non-redundant unigenes. RNA-seq results showed that 52 DEGs involved in lignin biosynthesis may be responsible for S. portulacastrum salt tolerance. Furthermore, 130 SDMs were identified, and the salt response could be attributed to the p-coumaryl alcohol-rich in lignin biosynthesis. The co-expression network that was constructed after comparing the different salt treatment processes showed that the p-Coumaryl alcohol was linked to 30 DEGs. Herein, 8 structures genes, i.e., Sp4CL, SpCAD, SpCCR, SpCOMT, SpF5H, SpCYP73A, SpCCoAOMT, and SpC3'H were identified as significant factors in regulating lignin biosynthesis. Further investigation revealed that 64 putative transcription factors (TFs) may interact with the promoters of the above-mentioned genes. Together, the data revealed a potential regulatory network comprising important genes, putative TFs, and metabolites involved in the lignin biosynthesis of S. portulacastrum roots under salt stress, which could serve as a rich useful genetic resource for breeding excellent salt-tolerant plants.
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Transcriptome diversity assessment of Gossypium arboreum (FDH228) leaves under control, drought and whitefly infestation using PacBio long reads. Gene 2023; 852:147065. [PMID: 36435508 DOI: 10.1016/j.gene.2022.147065] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/22/2022] [Revised: 11/08/2022] [Accepted: 11/18/2022] [Indexed: 11/25/2022]
Abstract
Alternative splicing (AS) and alternative polyadenylation (APA) are common mechanisms in eukaryotes to increase the complexity of transcriptomes and subsequently proteomes. Analysis of long reads transcriptomics data can result in the discovery of novel transcripts, splice sites, AS or APA events. Gossypium arboreum is an important cultivated cotton species and a putative contributor of the A sub-genome to the modern tetraploid cotton; and inherently tolerant to several biotic and abiotic stresses. Specifically, its variety 'FDH228' is considered to be an important resistance source. In this study, we sequenced the G. arboreum (var. FDH228) transcriptome using PacBio IsoSeq and illumina short read sequencing under three different conditions i.e. untreated/healthy, treated with biotic stress through whitefly infestation, and treated with abiotic stress via water deprivation, for the discovery and surveying of canonical and non-canonical AS, APA and transcript fusion events. We were able to obtain 15,419 unique transcripts from all samples representing 11,343 genes, out of which 10,832 were annotated and 520 were novel with respect to the published reference genome. These transcripts were grouped into different structural categories including 60 Antisense, 11,959 having a full-splice match, 999 with incomplete-splice match, 30 fusion transcripts, 177 genic, 479 intergenic, 771 novels in the catalog, and 944 Novel but not found in the catalog. Subsequently, randomly selected candidate transcripts were experimentally validated using qRT-PCR. Our comprehensive identification of canonical and non-canonical splicing events, and novel and fusion transcripts aids in the understanding of the resistance mechanisms for this specific germplasm.
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16
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Ma SH, He GQ, Navarro-Payá D, Santiago A, Cheng YZ, Jiao JB, Li HJ, Zuo DD, Sun HT, Pei MS, Yu YH, Matus JT, Guo DL. Global analysis of alternative splicing events based on long- and short-read RNA sequencing during grape berry development. Gene 2023; 852:147056. [PMID: 36414171 DOI: 10.1016/j.gene.2022.147056] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2022] [Revised: 11/09/2022] [Accepted: 11/14/2022] [Indexed: 11/21/2022]
Affiliation(s)
- Shuai-Hui Ma
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Guang-Qi He
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - David Navarro-Payá
- Institute for Integrative Systems Biology (I2SysBio), Universitat de València-CSIC, Paterna, Spain
| | - Antonio Santiago
- Institute for Integrative Systems Biology (I2SysBio), Universitat de València-CSIC, Paterna, Spain
| | - Yi-Zhe Cheng
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Jia-Bing Jiao
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Hui-Jie Li
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Ding-Ding Zuo
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Hao-Ting Sun
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Mao-Song Pei
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - Yi-He Yu
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China
| | - José Tomás Matus
- Institute for Integrative Systems Biology (I2SysBio), Universitat de València-CSIC, Paterna, Spain
| | - Da-Long Guo
- College of Horticulture and Plant Protection, Henan University of Science and Technology, Luoyang 471023, China; Henan Engineering Technology Research Center of Quality Regulation of Horticultural Plants, Luoyang 471023, China.
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17
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Bencurova E, Akash A, Dobson RC, Dandekar T. DNA storage-from natural biology to synthetic biology. Comput Struct Biotechnol J 2023; 21:1227-1235. [PMID: 36817961 PMCID: PMC9932295 DOI: 10.1016/j.csbj.2023.01.045] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2022] [Revised: 01/30/2023] [Accepted: 01/31/2023] [Indexed: 02/04/2023] Open
Abstract
Natural DNA storage allows cellular differentiation, evolution, the growth of our children and controls all our ecosystems. Here, we discuss the fundamental aspects of DNA storage and recent advances in this field, with special emphasis on natural processes and solutions that can be exploited. We point out new ways of efficient DNA and nucleotide storage that are inspired by nature. Within a few years DNA-based information storage may become an attractive and natural complementation to current electronic data storage systems. We discuss rapid and directed access (e.g. DNA elements such as promotors, enhancers), regulatory signals and modulation (e.g. lncRNA) as well as integrated high-density storage and processing modules (e.g. chromosomal territories). There is pragmatic DNA storage for use in biotechnology and human genetics. We examine DNA storage as an approach for synthetic biology (e.g. light-controlled nucleotide processing enzymes). The natural polymers of DNA and RNA offer much for direct storage operations (read-in, read-out, access control). The inbuilt parallelism (many molecules at many places working at the same time) is important for fast processing of information. Using biology concepts from chromosomal storage, nucleic acid processing as well as polymer material sciences such as electronical effects in enzymes, graphene, nanocellulose up to DNA macramé , DNA wires and DNA-based aptamer field effect transistors will open up new applications gradually replacing classical information storage methods in ever more areas over time (decades).
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Affiliation(s)
- Elena Bencurova
- Department of Bioinformatics, University of Würzburg, Würzburg, Germany
| | - Aman Akash
- Department of Bioinformatics, University of Würzburg, Würzburg, Germany
| | - Renwick C.J. Dobson
- Biomolecular Interaction Centre, University of Canterbury, Christchurch, New Zealand,Department of Biochemistry and Pharmacology, University of Melbourne, Melbourne, Australia
| | - Thomas Dandekar
- Department of Bioinformatics, University of Würzburg, Würzburg, Germany,Structural and Computational Biology, European Molecular Biology Laboratory, Heidelberg, Germany,Corresponding author at: Department of Bioinformatics, University of Würzburg, Würzburg, Germany.
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18
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Ruan H, Wang PC, Han L. Characterization of circular RNAs with advanced sequencing technologies in human complex diseases. WILEY INTERDISCIPLINARY REVIEWS. RNA 2023; 14:e1759. [PMID: 36164985 DOI: 10.1002/wrna.1759] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/28/2022] [Revised: 07/09/2022] [Accepted: 08/02/2022] [Indexed: 01/31/2023]
Abstract
Circular RNAs (circRNAs) are one category of non-coding RNAs that do not possess 5' caps and 3' free ends. Instead, they are derived in closed circle forms from pre-mRNAs by a non-canonical splicing mechanism named "back-splicing." CircRNAs were discovered four decades ago, initially called "scrambled exons." Compared to linear RNAs, the expression levels of circRNAs are considerably lower, and it is challenging to identify circRNAs specifically. Thus, the biological relevance of circRNAs has been underappreciated until the advancement of next generation sequencing (NGS) technology. The biological insights of circRNAs, such as their tissue-specific expression patterns, biogenesis factors, and functional effects in complex diseases, namely human cancers, have been extensively explored in the last decade. With the invention of the third generation sequencing (TGS) with longer sequencing reads and newly designed strategies to characterize full-length circRNAs, the panorama of circRNAs in human complex diseases could be further unveiled. In this review, we first introduce the history of circular RNA detection. Next, we describe widely adopted NGS-based methods and the recently established TGS-based approaches capable of characterizing circRNAs in full-length. We then summarize data resources and representative circRNA functional studies related to human complex diseases. In the last section, we reviewed computational tools and discuss the potential advantages of utilizing advanced sequencing approaches to a functional interpretation of full-length circRNAs in complex diseases. This article is categorized under: RNA Evolution and Genomics > Computational Analyses of RNA RNA in Disease and Development > RNA in Disease.
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Affiliation(s)
- Hang Ruan
- Institutes of Biology and Medical Sciences, Soochow University, Suzhou, China
| | - Peng-Cheng Wang
- Institutes of Biology and Medical Sciences, Soochow University, Suzhou, China
| | - Leng Han
- Center for Epigenetics and Disease Prevention, Institute of Biosciences and Technology, Texas A&M University, Houston, Texas, USA.,Department of Translational Medical Sciences, College of Medicine, Texas A&M University, Houston, Texas, USA
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19
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Yang Y, Sun Y, Wang Z, Yin M, Sun R, Xue L, Huang X, Wang C, Yan X. Full-length transcriptome and metabolite analysis reveal reticuline epimerase-independent pathways for benzylisoquinoline alkaloids biosynthesis in Sinomenium acutum. FRONTIERS IN PLANT SCIENCE 2022; 13:1086335. [PMID: 36605968 PMCID: PMC9808091 DOI: 10.3389/fpls.2022.1086335] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 11/01/2022] [Accepted: 12/01/2022] [Indexed: 06/17/2023]
Abstract
Benzylisoquinoline alkaloids (BIAs) are a large family of plant natural products with important pharmaceutical applications. Sinomenium acutum is a medicinal plant from the Menispermaceae family and has been used to treat rheumatoid arthritis for hundreds of years. Sinomenium acutum contains more than 50 BIAs, and sinomenine is a representative BIA from this plant. Sinomenine was found to have preventive and curative effects on opioid dependence. Despite the broad applications of S. acutum, investigation on the biosynthetic pathways of BIAs from S. acutum is limited. In this study, we comprehensively analyzed the transcriptome data and BIAs in the root, stem, leaf, and seed of S. acutum. Metabolic analysis showed a noticeable difference in BIA contents in different tissues. Based on the study of the full-length transcriptome, differentially expressed genes, and weighted gene co-expression network, we proposed the biosynthetic pathways for a few BIAs from S. acutum, such as sinomenine, magnoflorine, and tetrahydropalmatine, and screened candidate genes involved in these biosynthesis processes. Notably, the reticuline epimerase (REPI/STORR), which converts (S)-reticuline to (R)-reticuline and plays an essential role in morphine and codeine biosynthesis, was not found in the transcriptome data of S. acutum. Our results shed light on the biogenesis of the BIAs in S. acutum and may pave the way for the future development of this important medicinal plant.
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Affiliation(s)
- Yufan Yang
- State Key Laboratory of Component-based Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China
- Haihe Laboratory of Modern Chinese Medicine, Tianjin, China
| | - Ying Sun
- State Key Laboratory of Component-based Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China
- WuXi AppTec (Tianjin) Co., Ltd., Tianjin, China
| | - Zhaoxin Wang
- State Key Laboratory of Component-based Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China
- Haihe Laboratory of Modern Chinese Medicine, Tianjin, China
| | - Maojing Yin
- College of Pharmaceutical Engineering of Traditional Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China
- School of Medicine, Foshan University, Foshan, Guangdong, China
| | - Runze Sun
- Hunan Provincial Key Laboratory for Synthetic Biology of Traditional Chinese Medicine, Hunan University of Medicine, Huaihua, Hunan, China
| | - Lu Xue
- State Key Laboratory of Component-based Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China
- Haihe Laboratory of Modern Chinese Medicine, Tianjin, China
| | - Xueshuang Huang
- Hunan Provincial Key Laboratory for Synthetic Biology of Traditional Chinese Medicine, Hunan University of Medicine, Huaihua, Hunan, China
| | - Chunhua Wang
- School of Medicine, Foshan University, Foshan, Guangdong, China
| | - Xiaohui Yan
- State Key Laboratory of Component-based Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China
- Haihe Laboratory of Modern Chinese Medicine, Tianjin, China
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Athanasopoulou K, Daneva GN, Boti MA, Dimitroulis G, Adamopoulos PG, Scorilas A. The Transition from Cancer "omics" to "epi-omics" through Next- and Third-Generation Sequencing. LIFE (BASEL, SWITZERLAND) 2022; 12:life12122010. [PMID: 36556377 PMCID: PMC9785810 DOI: 10.3390/life12122010] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 11/02/2022] [Revised: 11/25/2022] [Accepted: 11/30/2022] [Indexed: 12/05/2022]
Abstract
Deciphering cancer etiopathogenesis has proven to be an especially challenging task since the mechanisms that drive tumor development and progression are far from simple. An astonishing amount of research has revealed a wide spectrum of defects, including genomic abnormalities, epigenomic alterations, disturbance of gene transcription, as well as post-translational protein modifications, which cooperatively promote carcinogenesis. These findings suggest that the adoption of a multidimensional approach can provide a much more precise and comprehensive picture of the tumor landscape, hence serving as a powerful tool in cancer research and precision oncology. The introduction of next- and third-generation sequencing technologies paved the way for the decoding of genetic information and the elucidation of cancer-related cellular compounds and mechanisms. In the present review, we discuss the current and emerging applications of both generations of sequencing technologies, also referred to as massive parallel sequencing (MPS), in the fields of cancer genomics, transcriptomics and proteomics, as well as in the progressing realms of epi-omics. Finally, we provide a brief insight into the expanding scope of sequencing applications in personalized cancer medicine and pharmacogenomics.
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21
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Hou L, Wang M, Zhu L, Ning M, Bi J, Du J, Kong X, Gu W, Meng Q. Full-length transcriptome sequencing and comparative transcriptome analysis of Eriocheir sinensis in response to infection by the microsporidian Hepatospora eriocheir. Front Cell Infect Microbiol 2022; 12:997574. [PMID: 36530442 PMCID: PMC9754153 DOI: 10.3389/fcimb.2022.997574] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/19/2022] [Accepted: 11/18/2022] [Indexed: 12/02/2022] Open
Abstract
As a new generation of high-throughput sequencing technology, PacBio Iso-Seq technology (Iso-Seq) provides a better alternative sequencing method for the acquisition of full-length unigenes. In this study, a total of 22.27 gigabyte (Gb) subread bases and 128,614 non-redundant unigenes (mean length: 2,324 bp) were obtained from six main tissues of Eriocheir sinensis including the heart, nerve, intestine, muscle, gills and hepatopancreas. In addition, 74,732 unigenes were mapped to at least one of the following databases: Non-Redundant Protein Sequence Database (NR), Gene Ontology (GO), Kyoto Encyclopaedia of Genes and Genomes (KEGG), KEGG Orthology (KO) and Protein family (Pfam). In addition, 6696 transcription factors (TFs), 28,458 long non-coding RNAs (lncRNAs) and 94,230 mRNA-miRNA pairs were identified. Hepatospora eriocheir is the primary pathogen of E. sinensis and can cause hepatopancreatic necrosis disease (HPND); the intestine is the main target tissue. Here, we attempted to identify the key genes related to H. eriocheir infection in the intestines of E. sinensis. By combining Iso-Seq and Illumina RNA-seq analysis, we identified a total of 12,708 differentially expressed unigenes (DEUs; 6,696 upregulated and 6,012 downregulated) in the crab intestine following infection with H. eriocheir. Based on the biological analysis of these DEUs, several key processes were identified, including energy metabolism-related pathways, cell apoptosis and innate immune-related pathways. Twelve selected genes from these DEUs were subsequently verified by quantitative real-time PCR (qRT-PCR) analysis. Our findings enhance our understanding of the E. sinensis transcriptome and the specific association between E. sinensis and H. eriocheir infection.
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Affiliation(s)
- Libo Hou
- Engineering Lab of Henan Province for Aquatic Animal Disease Control, College of Fisheries, Henan Normal University, Xinxiang, China
| | - Mengdi Wang
- Engineering Lab of Henan Province for Aquatic Animal Disease Control, College of Fisheries, Henan Normal University, Xinxiang, China
| | - Lei Zhu
- Engineering Lab of Henan Province for Aquatic Animal Disease Control, College of Fisheries, Henan Normal University, Xinxiang, China
| | - Mingxiao Ning
- Institution of Quality Standard and Testing Technology for Agro-product, Shandong Academy of Agricultural Science, Jinan, Shandong, China
| | - Jingxiu Bi
- Institution of Quality Standard and Testing Technology for Agro-product, Shandong Academy of Agricultural Science, Jinan, Shandong, China
| | - Jie Du
- Animal Husbandry and Veterinary College, Jiangsu Vocational College of Agriculture and Forestry, Jurong, Jiangsu, China
| | - Xianghui Kong
- Engineering Lab of Henan Province for Aquatic Animal Disease Control, College of Fisheries, Henan Normal University, Xinxiang, China
| | - Wei Gu
- Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Marine Science and Engineering, Nanjing Normal University, Nanjing, Jiangsu, China
| | - Qingguo Meng
- Jiangsu Key Laboratory for Aquatic Crustacean Diseases, College of Marine Science and Engineering, Nanjing Normal University, Nanjing, Jiangsu, China,*Correspondence: Qingguo Meng,
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Zhou T, He Y, Zeng X, Cai B, Qu S, Wang S. Comparative Analysis of Alternative Splicing in Two Contrasting Apple Cultivars Defense against Alternaria alternata Apple Pathotype Infection. Int J Mol Sci 2022; 23:ijms232214202. [PMID: 36430679 PMCID: PMC9693243 DOI: 10.3390/ijms232214202] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2022] [Revised: 11/03/2022] [Accepted: 11/14/2022] [Indexed: 11/19/2022] Open
Abstract
Alternaria blotch disease, caused by the Alternaria alternata apple pathotype (A. alternata AP), is one of the most serious fungal diseases in apples. Alternative splicing (AS), one of the pivotal post-transcriptional regulatory mechanisms, plays essential roles in various disease resistance responses. Here, we performed RNA-Seq for two apple cultivars (resistant cultivar 'Jonathan' (J) and susceptible cultivar 'Starking Delicious' (SD)) infected by A. alternata AP to further investigate their AS divergence. In total, 1454, 1780, 1367 and 1698 specifically regulated differential alternative splicing (DAS) events were detected in J36, J72, SD36 and SD72 groups, respectively. Retained intron (RI) was the dominant AS pattern. Conformably, 642, 764, 585 and 742 uniquely regulated differentially spliced genes (DSGs) were found during A. alternata AP infection. Comparative analysis of AS genes in differential splicing and expression levels suggested that only a small proportion of DSGs overlapped with differentially expressed genes (DEGs). Gene ontology (GO) enrichment analysis demonstrated that the DSGs were significantly enriched at multiple levels of gene expression regulation. Briefly, the specific AS was triggered in apple defense against A. alternata AP. Therefore, this study facilitates our understanding on the roles of AS regulation in response to A. alternata AP infection in apples.
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Single-molecule real-time sequencing of the full-length transcriptome of Halophila beccarii. Sci Rep 2022; 12:16444. [PMID: 36180578 PMCID: PMC9525579 DOI: 10.1038/s41598-022-20988-w] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2022] [Accepted: 09/21/2022] [Indexed: 11/24/2022] Open
Abstract
Ecologically, Halophila beccarii Asch. is considered as a colonizing or a pioneer seagrass species and a “tiny but mighty” seagrass species, since it may recover quickly from disturbance generally. The use of transcriptome technology can provide a better understanding of the physiological processes of seagrasses. To date, little is known about the genome and transcriptome information of H. beccarii. In this study, we used single molecule real-time (SMRT) sequencing to obtain full-length transcriptome data and characterize the transcriptome structure. A total of 11,773 of the 15,348 transcripts were successfully annotated in seven databases. In addition, 1573 long non-coding RNAs, 8402 simple sequence repeats and 2567 transcription factors were predicted in all the transcripts. A GO analysis showed that 5843 transcripts were divided into three categories, including biological process (BP), cellular component (CC) and molecular function (MF). In these three categories, metabolic process (1603 transcripts), protein-containing complex (515 transcripts) and binding (3233 transcripts) were the primary terms in BP, CC, and MF, respectively. The major types of transcription factors were involved in MYB-related and NF-YB families. To the best of our knowledge, this is the first report of the transcriptome of H. beccarii using SMRT sequencing technology.
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Srikakulam N, Sridevi G, Pandi G. High-quality reference transcriptome construction improves RNA-seq quantification in Oryza sativa indica. Front Genet 2022; 13:995072. [PMID: 36246658 PMCID: PMC9558114 DOI: 10.3389/fgene.2022.995072] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2022] [Accepted: 09/02/2022] [Indexed: 11/13/2022] Open
Abstract
The Reference Transcriptomic Dataset (RTD) is an accurate and comprehensive collection of transcripts originating from a given organism. It holds the key to precise transcript quantification and downstream analysis of differential expressions and regulations. Currently, transcriptome annotations for most crop plants are far from complete. For example, Oryza sativa indica (O. sativa indica) is reported to have 40,759 transcripts in the Ensembl database without alternative transcript isoforms and alternative splicing (AS) events. To generate a high-quality RTD, we conducted RNA sequencing of rice leaf samples collected at various time points during Rhizoctonia solani infection. The obtained reads were analyzed by adopting the recently developed computational analysis pipeline to assemble the RTD with increased transcript and AS diversity for O. sativa indica (IndicaRTD). After stringent quality filtering, the newly constructed transcriptome annotation was comprised of 122,968 non-redundant transcripts from 53,695 genes. This study identified many novel transcripts compared to Ensembl deposited data that are important for regulating molecular and physiological processes in the plant system. Currently, the assembled IndicaRTD must allow fast quantification of transcript and gene expression with high precision.
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Affiliation(s)
- Nagesh Srikakulam
- Laboratory of RNA Biology and Epigenomics, Department of Plant Biotechnology, School of Biotechnology, Madurai Kamaraj University, Madurai, India
- *Correspondence: Nagesh Srikakulam, ; Gopal Pandi,
| | - Ganapathi Sridevi
- Department of Plant Biotechnology, School of Biotechnology, Madurai Kamaraj University, Madurai, India
| | - Gopal Pandi
- Laboratory of RNA Biology and Epigenomics, Department of Plant Biotechnology, School of Biotechnology, Madurai Kamaraj University, Madurai, India
- *Correspondence: Nagesh Srikakulam, ; Gopal Pandi,
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Learning and memory impairment and transcriptomic profile in hippocampus of offspring after maternal fructose exposure during gestation and lactation. Food Chem Toxicol 2022; 169:113394. [PMID: 36049592 DOI: 10.1016/j.fct.2022.113394] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2022] [Revised: 08/06/2022] [Accepted: 08/24/2022] [Indexed: 11/22/2022]
Abstract
Increased fructose intake is a global issue, especially in mothers. Maternal fructose exposure during gestation and lactation can affect learning and memory in offspring; however, the detailed mechanism is still unknown. The hippocampus is a mind locale liable for learning and memory. Here, we established a maternal high-fructose diet model by administering 13% and 40% fructose water, applied the Morris Water Maze test on postnatal day 60 offspring, and performed full-length RNA sequencing using the Oxford Nanopore Technologies platform to explore the changes in gene expression in the hippocampus. The results showed that learning and memory in offspring were negatively affected. Compared with the control group, 369 differentially expressed transcripts (DETs) were identified in the 13% fructose group, and 501 DETs were identified in the 40% fructose group. Gene Ontology enriched term and Kyoto Encyclopedia of Genes and Genomes enriched pathway analyses identified several terms and pathways related to brain development and cognitive function. Furthermore, we confirmed that the Wnt/β-catenin signaling pathway was down-regulated and neuron degeneration was enhanced. In summary, our results indicate that maternal fructose exposure during gestation and lactation can impair learning and memory in offspring and affect brain function at the transcriptome level.
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Yang F, Lv G. Characterization of the gene expression profile response to drought stress in Haloxylon using PacBio single-molecule real-time and Illumina sequencing. FRONTIERS IN PLANT SCIENCE 2022; 13:981029. [PMID: 36051288 PMCID: PMC9424927 DOI: 10.3389/fpls.2022.981029] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 06/29/2022] [Accepted: 07/22/2022] [Indexed: 06/15/2023]
Abstract
Haloxylon ammodendron and Haloxylon persicum are important drought-tolerant plants in northwest China. The whole-genome sequencing of H. ammodendron and H. persicum grown in their natural environment is incomplete, and their transcriptional regulatory network in response to drought environment remains unclear. To reveal the transcriptional responses of H. ammodendron and H. persicum to an arid environment, we performed single-molecule real-time (SMRT) and Illumina RNA sequencing. In total, 20,246,576 and 908,053 subreads and 435,938 and 210,334 circular consensus sequencing (CCS) reads were identified by SMRT sequencing of H. ammodendron and H. persicum, and 15,238 and 10,135 unigenes, respectively, were successfully obtained. In addition, 9,794 and 7,330 simple sequence repeats (SSRs) and 838 and 71 long non-coding RNAs were identified. In an arid environment, the growth of H. ammodendron was restricted; plant height decreased significantly; basal and branch diameters became thinner and hydrogen peroxide (H2O2) content and peroxidase (POD) activity were increased. Under dry and wet conditions, 11,803 and 15,217 differentially expressed genes (DEGs) were identified in H. ammodendron and H. persicum, respectively. There were 319 and 415 DEGs in the signal transduction pathways related to drought stress signal perception and transmission, including the Ca2+ signal pathway, the ABA signal pathway, and the MAPK signal cascade. In addition, 217 transcription factors (TFs) and 398 TFs of H. ammodendron and H. persicum were differentially expressed, including FAR1, MYB, and AP2/ERF. Bioinformatic analysis showed that under drought stress, the expression patterns of genes related to active oxygen [reactive oxygen species (ROS)] scavenging, functional proteins, lignin biosynthesis, and glucose metabolism pathways were altered. Thisis the first full-length transcriptome report concerning the responses of H. ammodendron and H. persicum to drought stress. The results provide a foundation for further study of the adaptation to drought stress. The full-length transcriptome can be used in genetic engineering research.
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Affiliation(s)
- Fang Yang
- School of Ecology and Environment, Xinjiang University, Ürümqi, China
- Key Laboratory of Oasis Ecology, Ministry of Education, Ürümqi, China
- Xinjiang Jinghe Observation and Research Station of Temperate Desert Ecosystem, Ministry of Education, Ürümqi, China
| | - Guanghui Lv
- School of Ecology and Environment, Xinjiang University, Ürümqi, China
- Key Laboratory of Oasis Ecology, Ministry of Education, Ürümqi, China
- Xinjiang Jinghe Observation and Research Station of Temperate Desert Ecosystem, Ministry of Education, Ürümqi, China
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27
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Xiong J, Tang X, Wei M, Yu W. Comparative full-length transcriptome analysis by Oxford Nanopore Technologies reveals genes involved in anthocyanin accumulation in storage roots of sweet potatoes ( Ipomoea batatas L.). PeerJ 2022; 10:e13688. [PMID: 35846886 PMCID: PMC9285475 DOI: 10.7717/peerj.13688] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/19/2021] [Accepted: 06/16/2022] [Indexed: 01/17/2023] Open
Abstract
Background Storage roots of sweet potatoes (Ipomoea batatas L.) with different colors vary in anthocyanin content, indicating different economically agronomic trait. As the newest DNA/RNA sequencing technology, Oxford Nanopore Technologies (ONT) have been applied in rapid transcriptome sequencing for investigation of genes related to nutrient metabolism. At present, few reports concern full-length transcriptome analysis based on ONT for study on the molecular mechanism of anthocyanin accumulation leading to color change of tuberous roots of sweet potato cultivars. Results The storage roots of purple-fleshed sweet potato (PFSP) and white-fleshed sweet potato (WFSP) at different developmental stages were subjected to anthocyanin content comparison by UV-visible spectroscopy as well as transcriptome analysis at ONT MinION platform. UV-visible spectrophotometric measurements demonstrated the anthocyanin content of PFSP was much higher than that of WFSP. ONT RNA-Seq results showed each sample generated average 2.75 GB clean data with Full-Length Percentage (FL%) over 70% and the length of N50 ranged from 1,192 to 1,395 bp, indicating reliable data for transcriptome analysis. Subsequent analysis illustrated intron retention was the most prominent splicing event present in the resulting transcripts. As compared PFSP with WFSP at the relative developmental stages with the highest (PH vs. WH) and the lowest (PL vs. WL) anthocyanin content, 282 and 216 genes were up-regulated and two and 11 genes were down-regulated respectively. The differential expression genes involved in flavonoid biosynthesis pathway include CCoAOMT, PpLDOX, DFR, Cytochrome P450, CHI, and CHS. The genes encoding oxygenase superfamily were significantly up-regulated when compared PFSP with WFSP at the relative developmental stages. Conclusions Comparative full-length transcriptome analysis based on ONT serves as an effective approach to detect the differences in anthocyanin accumulation in the storage roots of different sweet potato cultivars at transcript level, with noting that some key genes can now be closely related to flavonoids biosynthesis. This study helps to improve understanding of molecular mechanism for anthocyanin accumulation in sweet potatoes and also provides a theoretical basis for high-quality sweet potato breeding.
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Affiliation(s)
- Jun Xiong
- Agricultural College, Guangxi University, Nanning, China,Cash Crops Research Institute, Guangxi Academy of Agricultural Sciences, Nanning, China
| | - Xiuhua Tang
- Cash Crops Research Institute, Guangxi Academy of Agricultural Sciences, Nanning, China
| | - Minzheng Wei
- Cash Crops Research Institute, Guangxi Academy of Agricultural Sciences, Nanning, China
| | - Wenjin Yu
- Agricultural College, Guangxi University, Nanning, China
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28
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Zhang R, Kuo R, Coulter M, Calixto CPG, Entizne JC, Guo W, Marquez Y, Milne L, Riegler S, Matsui A, Tanaka M, Harvey S, Gao Y, Wießner-Kroh T, Paniagua A, Crespi M, Denby K, Hur AB, Huq E, Jantsch M, Jarmolowski A, Koester T, Laubinger S, Li QQ, Gu L, Seki M, Staiger D, Sunkar R, Szweykowska-Kulinska Z, Tu SL, Wachter A, Waugh R, Xiong L, Zhang XN, Conesa A, Reddy ASN, Barta A, Kalyna M, Brown JWS. A high-resolution single-molecule sequencing-based Arabidopsis transcriptome using novel methods of Iso-seq analysis. Genome Biol 2022; 23:149. [PMID: 35799267 PMCID: PMC9264592 DOI: 10.1186/s13059-022-02711-0] [Citation(s) in RCA: 44] [Impact Index Per Article: 14.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2021] [Accepted: 06/15/2022] [Indexed: 12/15/2022] Open
Abstract
BACKGROUND Accurate and comprehensive annotation of transcript sequences is essential for transcript quantification and differential gene and transcript expression analysis. Single-molecule long-read sequencing technologies provide improved integrity of transcript structures including alternative splicing, and transcription start and polyadenylation sites. However, accuracy is significantly affected by sequencing errors, mRNA degradation, or incomplete cDNA synthesis. RESULTS We present a new and comprehensive Arabidopsis thaliana Reference Transcript Dataset 3 (AtRTD3). AtRTD3 contains over 169,000 transcripts-twice that of the best current Arabidopsis transcriptome and including over 1500 novel genes. Seventy-eight percent of transcripts are from Iso-seq with accurately defined splice junctions and transcription start and end sites. We develop novel methods to determine splice junctions and transcription start and end sites accurately. Mismatch profiles around splice junctions provide a powerful feature to distinguish correct splice junctions and remove false splice junctions. Stratified approaches identify high-confidence transcription start and end sites and remove fragmentary transcripts due to degradation. AtRTD3 is a major improvement over existing transcriptomes as demonstrated by analysis of an Arabidopsis cold response RNA-seq time-series. AtRTD3 provides higher resolution of transcript expression profiling and identifies cold-induced differential transcription start and polyadenylation site usage. CONCLUSIONS AtRTD3 is the most comprehensive Arabidopsis transcriptome currently. It improves the precision of differential gene and transcript expression, differential alternative splicing, and transcription start/end site usage analysis from RNA-seq data. The novel methods for identifying accurate splice junctions and transcription start/end sites are widely applicable and will improve single-molecule sequencing analysis from any species.
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Affiliation(s)
- Runxuan Zhang
- Information and Computational Sciences, James Hutton Institute, Dundee, DD2 5DA, Scotland, UK.
| | - Richard Kuo
- The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Midlothian, EH25 9RG, UK
| | - Max Coulter
- Plant Sciences Division, School of Life Sciences, University of Dundee at The James Hutton Institute, Invergowrie, Dundee, DD2 5DA, Scotland, UK
| | - Cristiane P G Calixto
- Plant Sciences Division, School of Life Sciences, University of Dundee at The James Hutton Institute, Invergowrie, Dundee, DD2 5DA, Scotland, UK
- Present address: Institute of Biosciences, University of São Paulo, São Paulo, 05508-090, Brazil
| | - Juan Carlos Entizne
- Plant Sciences Division, School of Life Sciences, University of Dundee at The James Hutton Institute, Invergowrie, Dundee, DD2 5DA, Scotland, UK
| | - Wenbin Guo
- Information and Computational Sciences, James Hutton Institute, Dundee, DD2 5DA, Scotland, UK
| | - Yamile Marquez
- Centre for Genomic Regulation, C/ Dr. Aiguader 88, 08003, Barcelona, Spain
| | - Linda Milne
- Information and Computational Sciences, James Hutton Institute, Dundee, DD2 5DA, Scotland, UK
| | - Stefan Riegler
- Institute of Molecular Plant Biology, Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences (BOKU), Muthgasse 18, 1190, Vienna, Austria
- Present address: Institute of Science and Technology Austria, Am Campus 1, 3400, Klosterneuburg, Austria
| | - Akihiro Matsui
- Plant Genomic Network Research Team, RIKEN Center for Sustainable Resource Science, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa, 230-0045, Japan
| | - Maho Tanaka
- Plant Genomic Network Research Team, RIKEN Center for Sustainable Resource Science, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa, 230-0045, Japan
| | - Sarah Harvey
- Centre for Novel Agricultural Products (CNAP), Department of Biology, University of York Wentworth Way, York, YO10 5DD, UK
| | - Yubang Gao
- College of Forestry, Fujian Agriculture and Forestry University, Fuzhou, 350002, China
| | - Theresa Wießner-Kroh
- Center for Plant Molecular Biology (ZMBP), University of Tübingen, Auf der Morgenstelle 32, 72076, Tübingen, Germany
| | - Alejandro Paniagua
- Institute for Integrative Systems Biology (CSIC-UV), Spanish National Research Council, Paterna, Valencia, Spain
| | - Martin Crespi
- French National Centre for Scientific Research | CNRS INRAE-Universities of Paris Saclay and Paris, Institute of Plant Sciences Paris Saclay IPS2, Rue de Noetzlin, 91192, Gif sur Yvette, France
| | - Katherine Denby
- Centre for Novel Agricultural Products (CNAP), Department of Biology, University of York Wentworth Way, York, YO10 5DD, UK
| | - Asa Ben Hur
- Department of Computer Science, Colorado State University, 1873 Campus Delivery, Fort Collins, CO, 80523-1873, USA
| | - Enamul Huq
- Department of Molecular Biosciences, University of Texas at Austin, 100 East 24th St., Austin, TX, 78712-1095, USA
| | - Michael Jantsch
- Department of Cell and Developmental Biology, Center for Anatomy and Cell Biology, Medical University of Vienna, Schwarzspanierstrasse 17 A-1090, Vienna, Austria
| | - Artur Jarmolowski
- Department of Gene Expression, Adam Mickiewicz University, Poznań, Poland
| | - Tino Koester
- RNA Biology and Molecular Physiology, Faculty for Biology, Bielefeld University, Universitaetsstrasse 25, 33615, Bielefeld, Germany
| | - Sascha Laubinger
- Institut für Biologie und Umweltwissenschaften (IBU), Carl von Ossietzky Universität Oldenburg, Carl von Ossietzky-Str. 9-11, 26111, Oldenburg, Germany
- Institute of Biology, Department of Genetics, Martin Luther University Halle-Wittenberg, Halle (Saale), Germany
| | - Qingshun Quinn Li
- Graduate College of Biomedical Sciences, Western University of Health Sciences, Pomona, CA, 91766, USA
- Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystems, College of the Environment and Ecology, Xiamen University, Xiamen, 361102, Fujian, China
| | - Lianfeng Gu
- College of Forestry, Fujian Agriculture and Forestry University, Fuzhou, 350002, China
| | - Motoaki Seki
- Plant Genomic Network Research Team, RIKEN Center for Sustainable Resource Science, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa, 230-0045, Japan
| | - Dorothee Staiger
- RNA Biology and Molecular Physiology, Faculty for Biology, Bielefeld University, Universitaetsstrasse 25, 33615, Bielefeld, Germany
| | - Ramanjulu Sunkar
- Department of Biochemistry and Molecular Biology, Oklahoma State University, Stillwater, OK, 74078, USA
| | | | - Shih-Long Tu
- Institute of Plant and Microbial Biology, Academia Sinica, Taipei, Taiwan
| | - Andreas Wachter
- Center for Plant Molecular Biology (ZMBP), University of Tübingen, Auf der Morgenstelle 32, 72076, Tübingen, Germany
- Present address: Institute for Molecular Physiology, Johannes Gutenberg University Mainz, Hanns-Dieter-Hüsch-Weg 17, 55128, Mainz, Germany
| | - Robbie Waugh
- Cell and Molecular Sciences, James Hutton Institute, Dundee, DD2 5DA, Scotland, UK
| | - Liming Xiong
- Department of Biology, Hong Kong Baptist University, Hong Kong, China
| | - Xiao-Ning Zhang
- Biology Department, School of Arts and Sciences, St. Bonaventure University, 3261 West State Road, St. Bonaventure, NY, 14778, USA
| | - Ana Conesa
- Institute for Integrative Systems Biology (CSIC-UV), Spanish National Research Council, Paterna, Valencia, Spain
| | - Anireddy S N Reddy
- Department of Biology and Program in Cell and Molecular Biology, Colorado State University, Fort Collins, CO, 80523, USA
| | - Andrea Barta
- Max F. Perutz Laboratories, Medical University of Vienna, Center of Medical Biochemistry, Dr.-Bohr-Gasse 9/3, A-1030, Vienna, Austria
| | - Maria Kalyna
- Institute of Molecular Plant Biology, Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences (BOKU), Muthgasse 18, 1190, Vienna, Austria
| | - John W S Brown
- Plant Sciences Division, School of Life Sciences, University of Dundee at The James Hutton Institute, Invergowrie, Dundee, DD2 5DA, Scotland, UK
- Cell and Molecular Sciences, James Hutton Institute, Dundee, DD2 5DA, Scotland, UK
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Lee KC, Lee HT, Jeong HH, Park JH, Kim YC, Lee JH, Kim JK. The splicing factor 1-FLOWERING LOCUS M module spatially regulates temperature-dependent flowering by modulating FLOWERING LOCUS T and LEAFY expression. PLANT CELL REPORTS 2022; 41:1603-1612. [PMID: 35589978 DOI: 10.1007/s00299-022-02881-y] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/05/2022] [Accepted: 04/28/2022] [Indexed: 06/15/2023]
Abstract
The AtSF1-FLM module spatially controls temperature-dependent flowering by negatively regulating the expression of FT and LFY in the leaf and shoot apex, respectively. Alternative splicing mediated by various splicing factors is important for the regulation of plant growth and development. Our recent reports have shown that a temperature-dependent interaction between Arabidopsis thaliana splicing factor 1 (AtSF1) and FLOWERING LOCUS M (FLM) pre-mRNA introns controls the differential production of FLM-β transcripts at different temperatures, eventually resulting in temperature-responsive flowering. However, the molecular and genetic interactions between the AtSF1-FLM module and floral activator genes remain unknown. Here, we aimed to identify the interactions among AtSF1, FLM, FLOWERING LOCUS T (FT), and LEAFY (LFY) by performing molecular and genetic analyses. FT and TWIN SISTER OF FT (TSF) expression in atsf1-2 mutants significantly increased in the morning and middle of the night at 16 and 23 °C, respectively, under long-day conditions. In addition, ft mutation suppressed the early flowering of atsf1-2 and atsf1-2 flm-3 mutants and masked the temperature response of atsf1-2 flm-3 mutants, suggesting that FT is a downstream target gene of the AtSF1-FLM module. LFY expression significantly increased in the diurnal samples of atsf1-2 mutants and in the shoot apex regions of atsf1-2 ft-10 mutants at different temperatures. The chromatin immunoprecipitation (ChIP) assay revealed that FLM directly binds to the genomic regions of LFY but not of APETALA1 (AP1). Moreover, lfy mutation suppressed the early flowering of flm-3 mutants, suggesting that LFY is another target of the AtSF1-FLM module. Our results reveal that the AtSF1-FLM module spatially modulates temperature-dependent flowering by regulating FT and LFY expressions.
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Affiliation(s)
- Keh Chien Lee
- Division of Life Sciences, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul, 02841, Republic of Korea
| | - Hee Tae Lee
- Division of Life Sciences, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul, 02841, Republic of Korea
| | - Hwa Hyun Jeong
- Division of Life Sciences, Jeonbuk National University, 567 Baekje-daero, Deokjin-gu, Jeonju, Jeollabuk-do, 54896, Republic of Korea
| | - Jae-Hyeok Park
- Division of Life Sciences, Jeonbuk National University, 567 Baekje-daero, Deokjin-gu, Jeonju, Jeollabuk-do, 54896, Republic of Korea
| | - Young-Cheon Kim
- Division of Life Sciences, Jeonbuk National University, 567 Baekje-daero, Deokjin-gu, Jeonju, Jeollabuk-do, 54896, Republic of Korea
| | - Jeong Hwan Lee
- Division of Life Sciences, Jeonbuk National University, 567 Baekje-daero, Deokjin-gu, Jeonju, Jeollabuk-do, 54896, Republic of Korea.
| | - Jeong-Kook Kim
- Division of Life Sciences, Korea University, 145 Anam-ro, Seongbuk-gu, Seoul, 02841, Republic of Korea.
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Hasan S, Huang L, Liu Q, Perlo V, O’Keeffe A, Margarido GRA, Furtado A, Henry RJ. The Long Read Transcriptome of Rice (Oryza sativa ssp. japonica var. Nipponbare) Reveals Novel Transcripts. RICE (NEW YORK, N.Y.) 2022; 15:29. [PMID: 35689714 PMCID: PMC9188635 DOI: 10.1186/s12284-022-00577-1] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/14/2022] [Accepted: 05/26/2022] [Indexed: 05/08/2023]
Abstract
BACKGROUND High-throughput next-generation sequencing technologies offer a powerful approach to characterizing the transcriptomes of plants. Long read sequencing has been shown to support the discovery of novel isoforms of transcripts. This approach enables the generation of full-length sequences revealing splice variants that may be important in regulating gene action. Investigation of the diversity of transcripts in the rice transcriptome including splice variants was conducted using PacBio long-read sequence data to improve the annotation of the rice genome. RESULTS A cDNA library was prepared from RNA extracted from leaves, roots, seeds, inflorescences, and panicles of O. sativa ssp. japonica var Nipponbare and sequenced on a PacBio Sequel platform. This produced 346,190 non-redundant full-length non-chimeric reads (FLNC) resulting in 33,504 high-quality transcripts. Half of the transcripts were multi-exonic and entirely matched with the reference transcripts. However, 14,874 novel isoforms were also identified resulting predominantly from intron retention and at least one novel splice site. Intron retention was the prevalent alternative splicing event and exon skipping was the least observed. Of 73,659 splice junctions, 12,755 (17%) represented novel splice junctions with canonical and non-canonical intron boundaries. The complexity of the transcriptome was examined in detail for 19 starch synthesis-related genes, defining 276 spliced isoforms of which 94 splice variants were novel. CONCLUSION The data reveal the great complexity of the rice transcriptome. The novel transcripts provide new insights that may be a key input in future research to improve the annotation of the rice genome.
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Affiliation(s)
- Sharmin Hasan
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072 Australia
- Department of Botany, Jagannath University, Dhaka, 1100 Bangladesh
| | - Lichun Huang
- College of Agriculture, Yangzhou University, Jiangsu, 225009 China
| | - Qiaoquan Liu
- College of Agriculture, Yangzhou University, Jiangsu, 225009 China
| | - Virginie Perlo
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072 Australia
| | - Angela O’Keeffe
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072 Australia
| | - Gabriel Rodrigues Alves Margarido
- Departamento de Genética, Escola Superior de Agricultura “Luiz de Queiroz”, Universidade de São Paulo, Piracicaba, São Paulo 13418-900 Brazil
| | - Agnelo Furtado
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072 Australia
| | - Robert J. Henry
- Queensland Alliance for Agriculture and Food Innovation, University of Queensland, Brisbane, 4072 Australia
- ARC Centre of Excellence for Plant Success in Nature and Agriculture, University of Queensland, Brisbane, 4072 Australia
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Huang M, Jiang Y, Qin R, Jiang D, Chang D, Tian Z, Li C, Wang C. Full-Length Transcriptional Analysis of the Same Soybean Genotype With Compatible and Incompatible Reactions to Heterodera glycines Reveals Nematode Infection Activating Plant Defense Response. FRONTIERS IN PLANT SCIENCE 2022; 13:866322. [PMID: 35665156 PMCID: PMC9158574 DOI: 10.3389/fpls.2022.866322] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/31/2022] [Accepted: 03/22/2022] [Indexed: 06/04/2023]
Abstract
Full-length transcriptome sequencing with long reads is a powerful tool to analyze transcriptional and post-transcriptional events; however, it has not been applied on soybean (Glycine max). Here, a comparative full-length transcriptome analysis was performed on soybean genotype 09-138 infected with soybean cyst nematode (SCN, Heterodera glycines) race 4 (SCN4, incompatible reaction) and race 5 (SCN5, compatible reaction) using Oxford Nanopore Technology. Each of 9 full-length samples collected 8 days post inoculation with/without nematodes generated an average of 6.1 GB of clean data and a total of 65,038 transcript sequences. After redundant transcripts were removed, 1,117 novel genes and 41,096 novel transcripts were identified. By analyzing the sequence structure of the novel transcripts, a total of 28,759 complete open reading frame (ORF) sequences, 5,337 transcription factors, 288 long non-coding RNAs, and 40,090 novel transcripts with function annotation were predicted. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses of differentially expressed genes (DEGs) revealed that growth hormone, auxin-activated signaling pathway and multidimensional cell growth, and phenylpropanoid biosynthesis pathway were enriched by infection with both nematode races. More DEGs associated with stress response elements, plant-hormone signaling transduction pathway, and plant-pathogen interaction pathway with more upregulation were found in the incompatible reaction with SCN4 infection, and more DEGs with more upregulation involved in cell wall modification and carbohydrate bioprocess were detected in the compatible reaction with SCN5 infection when compared with each other. Among them, overlapping DEGs with a quantitative difference was triggered. The combination of protein-protein interaction with DEGs for the first time indicated that nematode infection activated the interactions between transcription factor WRKY and VQ (valine-glutamine motif) to contribute to soybean defense. The knowledge of the SCN-soybean interaction mechanism as a model will present more understanding of other plant-nematode interactions.
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Affiliation(s)
- Minghui Huang
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
| | - Ye Jiang
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
- Heilongjiang Academy of Agricultural Sciences, Daqing, China
| | - Ruifeng Qin
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
- Heilongjiang Academy of Agricultural Sciences, Daqing, China
| | - Dan Jiang
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
- Heilongjiang Academy of Agricultural Sciences, Daqing, China
| | - Doudou Chang
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
- Heilongjiang Academy of Agricultural Sciences, Daqing, China
| | - Zhongyan Tian
- Heilongjiang Academy of Agricultural Sciences, Daqing, China
| | - Chunjie Li
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
| | - Congli Wang
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Harbin, China
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Xu T, Yang X, Jia Y, Li Z, Tang G, Li X, Wang B, Wang T, Lin J, Guo L, Ye K. A global survey of the transcriptome of the opium poppy (Papaver somniferum) based on single-molecule long-read isoform sequencing. THE PLANT JOURNAL : FOR CELL AND MOLECULAR BIOLOGY 2022; 110:607-620. [PMID: 35092713 DOI: 10.1111/tpj.15689] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/13/2021] [Revised: 01/13/2022] [Accepted: 01/19/2022] [Indexed: 06/14/2023]
Affiliation(s)
- Tun Xu
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- MOE Key Lab for Intelligent Networks & Networks Security, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
| | - Xiaofei Yang
- MOE Key Lab for Intelligent Networks & Networks Security, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- School of Computer Science and Technology, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- Genome Institute, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, China
| | - Yanyan Jia
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
| | - Zihang Li
- School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, China
| | - Guangbo Tang
- School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, China
| | - Xiujuan Li
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
| | - Bo Wang
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
| | - Tingjie Wang
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
| | - Jiadong Lin
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- MOE Key Lab for Intelligent Networks & Networks Security, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- Faculty of Science, Leiden University, Leiden, The Netherlands
| | - Li Guo
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- MOE Key Lab for Intelligent Networks & Networks Security, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, China
| | - Kai Ye
- School of Automation Science and Engineering, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- MOE Key Lab for Intelligent Networks & Networks Security, Faculty of Electronic and Information Engineering, Xi'an Jiaotong University, Xi'an, China
- Genome Institute, The First Affiliated Hospital of Xi'an Jiaotong University, Xi'an, China
- School of Life Science and Technology, Xi'an Jiaotong University, Xi'an, China
- Faculty of Science, Leiden University, Leiden, The Netherlands
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He W, Zhang X, Lv P, Wang W, Wang J, He Y, Song Z, Cai D. Full-length transcriptome reconstruction reveals genetic differences in hybrids of Oryza sativa and Oryza punctata with different ploidy and genome compositions. BMC PLANT BIOLOGY 2022; 22:131. [PMID: 35313821 PMCID: PMC8935693 DOI: 10.1186/s12870-022-03502-2] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/07/2021] [Accepted: 03/01/2022] [Indexed: 07/07/2023]
Abstract
BACKGROUND Allopolyploid breeding is an efficient technique for improving the low seed setting rate of autotetraploids in plant breeding and one of the most promising breeding methods. However, there have been few comprehensive studies of the posttranscriptional mechanism in allopolyploids. RESULTS By crossing cultivated rice (Oryza sativa, genome AA) with wild rice (Oryza punctata, genome BB), we created hybrid rice lines with different ploidy and genome compositions [diploid hybrid F01 (AB), allotetraploid hybrid F02 (AABB) and F03 (AAAB)]. The genetic differences of the hybrids and the mechanism of allopolyploid breeding dominance were revealed through morphological and cytological observations and single molecule real-time sequencing techniques. The tissues and organs of allotetraploid hybrid F02 exhibited "gigantism" and the highest levels of fertility. The numbers of non-redundant transcripts, gene loci and new isoforms in the polyploid rice lines were higher and the isoform lengths greater than those of the diploid line. Moreover, alternative splicing (AS) events occurred twice as often in the polyploid rice lines than the diploid line. During these events, intron retention dominated. Furthermore, a large number of new genes and isoforms specific to the lines of different ploidy were discovered. CONCLUSIONS The results indicated that alternative polyadenylation (APA) and AS events contributed to the complexity and superiority of polyploids in the activity of translation regulators, nucleic acid binding transcription factor activities and the regulation of molecular function. Therefore, these APA and AS events in allopolyploid rice were found to play a role in regulation. Our study provides new germplasm for polyploid rice breeding and reveals complex regulatory mechanisms that may be related to heterosis and fertility.
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Affiliation(s)
- Wenting He
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
| | - Xianhua Zhang
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
- State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
| | - Pincang Lv
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
| | - Wei Wang
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
| | - Jie Wang
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
| | - Yuchi He
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
- State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China
| | - Zhaojian Song
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China.
- State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China.
| | - Detian Cai
- School of Life Sciences, Hubei University, Wuhan, 430062, People's Republic of China.
- Wuhan Polyploid Biotechnology Co., Ltd., Wuhan, 430345, People's Republic of China.
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Li D, Shao L, Xu T, Wang X, Zhang R, Zhang K, Xia Y, Zhang J. Hybrid RNA Sequencing Strategy for the Dynamic Transcriptomes of Winter Dormancy in an Evergreen Herbaceous Perennial, Iris japonica. Front Genet 2022; 13:841957. [PMID: 35368689 PMCID: PMC8965894 DOI: 10.3389/fgene.2022.841957] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2021] [Accepted: 02/11/2022] [Indexed: 11/30/2022] Open
Abstract
Japanese iris (Iris japonica) is a popular perennial ornamental that originated in China; it has a long display period and remains green outdoors throughout the year. winter dormancy characteristics contribute greatly to the evergreenness of herbaceous perennials. Thus, it is crucial to explore the mechanism of winter dormancy in this evergreen herbaceous perennial. Here, we used the hybrid RNA-seq strategy including single-molecule real-time (SMRT) and next-generation sequencing (NGS) technologies to generate large-scale Full-length transcripts to examine the shoot apical meristems of Japanese iris. A total of 10.57 Gb clean data for SMRT and over 142 Gb clean data for NGS were generated. Using hybrid error correction, 58,654 full-length transcripts were acquired and comprehensively analysed, and their expression levels were validated by real-time qPCR. This is the first full-length RNA-seq study in the Iris genus; our results provide a valuable resource and improve understanding of RNA processing in this genus, for which little genomic information is available as yet. In addition, our data will facilitate in-depth analyses of winter dormancy mechanisms in herbaceous perennials, especially evergreen monocotyledons.
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Affiliation(s)
| | | | | | | | | | | | - Yiping Xia
- *Correspondence: Jiaping Zhang, ; Yiping Xia,
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Sun M, Sun S, Mao C, Zhang H, Ou C, Jia Z, Wang Y, Ma W, Li M, Jia S, Mao P. Dynamic Responses of Antioxidant and Glyoxalase Systems to Seed Aging Based on Full-Length Transcriptome in Oat (Avena sativa L.). Antioxidants (Basel) 2022; 11:antiox11020395. [PMID: 35204277 PMCID: PMC8869221 DOI: 10.3390/antiox11020395] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/27/2022] [Revised: 02/12/2022] [Accepted: 02/13/2022] [Indexed: 12/20/2022] Open
Abstract
Seed aging is a major challenge for food security, agronomic production, and germplasm conservation, and reactive oxygen species (ROS) and methylglyoxal (MG) are highly involved in the aging process. However, the regulatory mechanisms controlling the abundance of ROS and MG are not well characterized. To characterize dynamic response of antioxidant and glyoxalase systems during seed aging, oat (Avena sativa L.) aged seeds with a range of germination percentages were used to explore physiological parameters, biochemical parameters and relevant gene expression. A reference transcriptome based on PacBio sequencing generated 67,184 non-redundant full-length transcripts, with 59,050 annotated. Subsequently, eleven seed samples were used to investigate the dynamic response of respiration, ROS and MG accumulation, antioxidant enzymes and glyoxalase activity, and associated genes expression. The 48 indicators with high correlation coefficients were divided into six major response patterns, and were used for placing eleven seed samples into four groups, i.e., non-aged (Group N), higher vigor (Group H), medium vigor (Group M), and lower vigor (Group L). Finally, we proposed a putative model for aging response and self-detoxification mechanisms based on the four groups representing different aging levels. In addition, the outcomes of the study suggested the dysfunction of antioxidant and glyoxalase system, and the accumulation of ROS and MG definitely contribute to oat seed aging.
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Guo XF, Zhou YL, Liu M, Wang ZW, Gui JF. Integrated application of Iso-seq and RNA-seq provides insights into unsynchronized growth in red swamp crayfish (Procambarus clarkii). AQUACULTURE REPORTS 2022; 22:101008. [DOI: 10.1016/j.aqrep.2022.101008] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/05/2025]
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Zhang Z, Guo J, Cai X, Li Y, Xi X, Lin R, Liang J, Wang X, Wu J. Improved Reference Genome Annotation of Brassica rapa by Pacific Biosciences RNA Sequencing. FRONTIERS IN PLANT SCIENCE 2022; 13:841618. [PMID: 35371168 PMCID: PMC8968949 DOI: 10.3389/fpls.2022.841618] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/22/2021] [Accepted: 02/17/2022] [Indexed: 05/05/2023]
Abstract
The species Brassica rapa includes several important vegetable crops. The draft reference genome of B. rapa ssp. pekinensis was completed in 2011, and it has since been updated twice. The pangenome with structural variations of 18 B. rapa accessions was published in 2021. Although extensive genomic analysis has been conducted on B. rapa, a comprehensive genome annotation including gene structure, alternative splicing (AS) events, and non-coding genes is still lacking. Therefore, we used the Pacific Biosciences (PacBio) single-molecular long-read technology to improve gene models and produced the annotated genome version 3.5. In total, we obtained 753,041 full-length non-chimeric (FLNC) reads and collapsed these into 92,810 non-redundant consensus isoforms, capturing 48% of the genes annotated in the B. rapa reference genome annotation v3.1. Based on the isoform data, we identified 830 novel protein-coding genes that were missed in previous genome annotations, defined the untranslated regions (UTRs) of 20,340 annotated genes and corrected 886 wrongly spliced genes. We also identified 28,564 AS events and 1,480 long non-coding RNAs (lncRNAs). We produced a relatively complete and high-quality reference transcriptome for B. rapa that can facilitate further functional genomic research.
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Olson AJ, Ware D. Ranked choice voting for representative transcripts with TRaCE. Bioinformatics 2021; 38:261-264. [PMID: 34297055 PMCID: PMC8696091 DOI: 10.1093/bioinformatics/btab542] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2020] [Revised: 06/09/2021] [Accepted: 07/22/2021] [Indexed: 02/03/2023] Open
Abstract
SUMMARY Genome sequencing projects annotate protein-coding gene models with multiple transcripts, aiming to represent all of the available transcript evidence. However, downstream analyses often operate on only one representative transcript per gene locus, sometimes known as the canonical transcript. To choose canonical transcripts, Transcript Ranking and Canonical Election (TRaCE) holds an 'election' in which a set of RNA-seq samples rank transcripts by annotation edit distance. These sample-specific votes are tallied along with other criteria such as protein length and InterPro domain coverage. The winner is selected as the canonical transcript, but the election proceeds through multiple rounds of voting to order all the transcripts by relevance. Based on the set of expression data provided, TRaCE can identify the most common isoforms from a broad expression atlas or prioritize alternative transcripts expressed in specific contexts. AVAILABILITY AND IMPLEMENTATION Transcript ranking code can be found on GitHub at {{https://github.com/warelab/TRaCE}}. SUPPLEMENTARY INFORMATION Supplementary data are available at Bioinformatics online.
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Affiliation(s)
- Andrew J Olson
- Plant Biology, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11768, USA
| | - Doreen Ware
- Plant Biology, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY 11768, USA
- USDA ARS Robert W. Holley Center for Agriculture and Health Cornell University, Ithaca, NY 14853, USA
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Molecular Analysis Uncovers the Mechanism of Fertility Restoration in Temperature-Sensitive Polima Cytoplasmic Male-Sterile Brassica napus. Int J Mol Sci 2021; 22:ijms222212450. [PMID: 34830333 PMCID: PMC8617660 DOI: 10.3390/ijms222212450] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2021] [Revised: 11/10/2021] [Accepted: 11/11/2021] [Indexed: 11/17/2022] Open
Abstract
Temperature-sensitive male sterility is a heritable agronomic trait affected by genotype-environment interactions. In rapeseed (Brassica napus), Polima (pol) temperature-sensitive cytoplasmic male sterility (TCMS) is commonly used for two-line breeding, as the fertility of pol TCMS lines can be partially restored at certain temperatures. However, little is known about the underlying molecular mechanism that controls fertility restoration. Therefore, we aimed to investigate the fertility conversion mechanism of the pol TCMS line at two different ambient temperatures (16 °C and 25 °C). Our results showed that the anthers developed and produced vigorous pollen at 16 °C but not at 25 °C. In addition, we identified a novel co-transcript of orf224-atp6 in the mitochondria that might lead to fertility conversion of the pol TCMS line. RNA-seq analysis showed that 1637 genes were significantly differentially expressed in the fertile flowers of 596-L when compared to the sterile flower of 1318 and 596-H. Detailed analysis revealed that differentially expressed genes were involved in temperature response, ROS accumulation, anther development, and mitochondrial function. Single-molecule long-read isoform sequencing combined with RNA sequencing revealed numerous genes produce alternative splicing transcripts at high temperatures. Here, we also found that alternative oxidase, type II NAD(P)H dehydrogenases, and transcription factor Hsfs might play a crucial role in male fertility under the low-temperature condition. RNA sequencing and bulked segregant analysis coupled with whole-genome sequencing identified the candidate genes involved in the post-transcriptional modification of orf224. Overall, our study described a putative mechanism of fertility restoration in a pol TCMS line controlled by ambient temperature that might help utilise TCMS in the two-line breeding of Brassica crops.
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Ebrahimie E, Rahimirad S, Tahsili M, Mohammadi-Dehcheshmeh M. Alternative RNA splicing in stem cells and cancer stem cells: Importance of transcript-based expression analysis. World J Stem Cells 2021; 13:1394-1416. [PMID: 34786151 PMCID: PMC8567453 DOI: 10.4252/wjsc.v13.i10.1394] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/18/2021] [Revised: 06/21/2021] [Accepted: 09/14/2021] [Indexed: 02/06/2023] Open
Abstract
Alternative ribonucleic acid (RNA) splicing can lead to the assembly of different protein isoforms with distinctive functions. The outcome of alternative splicing (AS) can result in a complete loss of function or the acquisition of new functions. There is a gap in knowledge of abnormal RNA splice variants promoting cancer stem cells (CSCs), and their prospective contribution in cancer progression. AS directly regulates the self-renewal features of stem cells (SCs) and stem-like cancer cells. Notably, octamer-binding transcription factor 4A spliced variant of octamer-binding transcription factor 4 contributes to maintaining stemness properties in both SCs and CSCs. The epithelial to mesenchymal transition pathway regulates the AS events in CSCs to maintain stemness. The alternative spliced variants of CSCs markers, including cluster of differentiation 44, aldehyde dehydrogenase, and doublecortin-like kinase, α6β1 integrin, have pivotal roles in increasing self-renewal properties and maintaining the pluripotency of CSCs. Various splicing analysis tools are considered in this study. LeafCutter software can be considered as the best tool for differential splicing analysis and identification of the type of splicing events. Additionally, LeafCutter can be used for efficient mapping splicing quantitative trait loci. Altogether, the accumulating evidence re-enforces the fact that gene and protein expression need to be investigated in parallel with alternative splice variants.
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Affiliation(s)
- Esmaeil Ebrahimie
- School of Animal and Veterinary Sciences, The University of Adelaide, Adelaide 5005, South Australia, Australia
- La Trobe Genomics Research Platform, School of Life Sciences, College of Science, Health and Engineering, La Trobe University, Melbourne 3086, Australia
- School of Biosciences, The University of Melbourne, Melbourne 3010, Australia,
| | - Samira Rahimirad
- Department of Medical Genetics, National Institute of Genetic Engineering and Biotechnology, Tehran 1497716316, Iran
- Division of Urology, Department of Surgery, McGill University and the Research Institute of the McGill University Health Centre, Montreal H4A 3J1, Quebec, Canada
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Yang J, Lv W, Shao L, Fu Y, Liu H, Yang C, Chen A, Xie X, Wang Z, Li C. PacBio and Illumina RNA Sequencing Identify Alternative Splicing Events in Response to Cold Stress in Two Poplar Species. FRONTIERS IN PLANT SCIENCE 2021; 12:737004. [PMID: 34691113 PMCID: PMC8529222 DOI: 10.3389/fpls.2021.737004] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/06/2021] [Accepted: 09/02/2021] [Indexed: 06/04/2023]
Abstract
In eukaryotes, alternative splicing (AS) is a crucial regulatory mechanism that modulates mRNA diversity and stability. The contribution of AS to stress is known in many species related to stress, but the posttranscriptional mechanism in poplar under cold stress is still unclear. Recent studies have utilized the advantages of single molecular real-time (SMRT) sequencing technology from Pacific Bioscience (PacBio) to identify full-length transcripts. We, therefore, used a combination of single-molecule long-read sequencing and Illumina RNA sequencing (RNA-Seq) for a global analysis of AS in two poplar species (Populus trichocarpa and P. ussuriensis) under cold stress. We further identified 1,261 AS events in P. trichocarpa and 2,101 in P. ussuriensis among which intron retention, with a frequency of more than 30%, was the most prominent type under cold stress. RNA-Seq data analysis and annotation revealed the importance of calcium, abscisic acid, and reactive oxygen species signaling in cold stress response. Besides, the low temperature rapidly induced multiple splicing factors, transcription factors, and differentially expressed genes through AS. In P. ussuriensis, there was a rapid occurrence of AS events, which provided a new insight into the complexity and regulation of AS during cold stress response in different poplar species for the first time.
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Affiliation(s)
- Jingli Yang
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Wanqiu Lv
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Liying Shao
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Yanrui Fu
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Haimei Liu
- Biology Group, Jiamusi No.1 High School, Jiamusi, China
| | - Chengjun Yang
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Aihua Chen
- Economic Forest Laboratory, Mudanjiang Branch of Heilongjiang Academy of Forestry, Mudanjiang, China
| | - Xieyu Xie
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Zhiwei Wang
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
| | - Chenghao Li
- State Key Laboratory of Forest Genetics and Tree Breeding, Northeast Forestry University, Harbin, China
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Mao L, Jin B, Chen L, Tian M, Ma R, Yin B, Zhang H, Guo J, Tang J, Chen T, Lai C, Cui G, Huang L. Functional identification of the terpene synthase family involved in diterpenoid alkaloids biosynthesis in Aconitum carmichaelii. Acta Pharm Sin B 2021; 11:3310-3321. [PMID: 34729318 PMCID: PMC8546855 DOI: 10.1016/j.apsb.2021.04.008] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/18/2021] [Revised: 03/22/2021] [Accepted: 04/02/2021] [Indexed: 02/07/2023] Open
Abstract
Aconitum carmichaelii is a high-value medicinal herb widely used across China, Japan, and other Asian countries. Aconitine-type diterpene alkaloids (DAs) are the characteristic compounds in Aconitum. Although six transcriptomes, based on short-read next generation sequencing technology, have been reported from the Aconitum species, the terpene synthase (TPS) corresponding to DAs biosynthesis remains unidentified. We apply a combination of Pacbio isoform sequencing and RNA sequencing to provide a comprehensive view of the A. carmichaelii transcriptome. Nineteen TPSs and five alternative splicing isoforms belonging to TPS-b, TPS-c, and TPS-e/f subfamilies were identified. In vitro enzyme reaction analysis functional identified two sesqui-TPSs and twelve diTPSs. Seven of the TPS-c subfamily genes reacted with GGPP to produce the intermediate ent-copalyl diphosphate. Five AcKSLs separately reacted with ent-CPP to produce ent-kaurene, ent-atiserene, and ent-13-epi-sandaracopimaradie: a new diterpene found in Aconitum. AcTPSs gene expression in conjunction DAs content analysis in different tissues validated that ent-CPP is the sole precursor to all DAs biosynthesis, with AcKSL1, AcKSL2s and AcKSL3-1 responsible for C20 atisine and napelline type DAs biosynthesis, respectively. These data clarified the molecular basis for the C20-DAs biosynthetic pathway in A. carmichaelii and pave the way for further exploration of C19-DAs biosynthesis in the Aconitum species.
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Affiliation(s)
- Liuying Mao
- College of Pharmacy, Shandong University of Chinese Medicine, Jinan 250355, China
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Baolong Jin
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Lingli Chen
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Mei Tian
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Rui Ma
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Biwei Yin
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Haiyan Zhang
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Juan Guo
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Jinfu Tang
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Tong Chen
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Changjiangsheng Lai
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Guanghong Cui
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
| | - Luqi Huang
- College of Pharmacy, Shandong University of Chinese Medicine, Jinan 250355, China
- State Key Laboratory of Dao-di Herbs, National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China
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Wang Y, Li X, Wang C, Gao L, Wu Y, Ni X, Sun J, Jiang J. Unveiling the transcriptomic complexity of Miscanthus sinensis using a combination of PacBio long read- and Illumina short read sequencing platforms. BMC Genomics 2021; 22:690. [PMID: 34551715 PMCID: PMC8459517 DOI: 10.1186/s12864-021-07971-x] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/15/2020] [Accepted: 09/03/2021] [Indexed: 11/10/2022] Open
Abstract
Background Miscanthus sinensis Andersson is a perennial grass that exhibits remarkable lignocellulose characteristics suitable for sustainable bioenergy production. However, knowledge of the genetic resources of this species is relatively limited, which considerably hampers further work on its biology and genetic improvement. Results In this study, through analyzing the transcriptome of mixed samples of leaves and stems using the latest PacBio Iso-Seq sequencing technology combined with Illumina HiSeq, we report the first full-length transcriptome dataset of M. sinensis with a total of 58.21 Gb clean data. An average of 15.75 Gb clean reads of each sample were obtained from the PacBio Iso-Seq system, which doubled the data size (6.68 Gb) obtained from the Illumina HiSeq platform. The integrated analyses of PacBio- and Illumina-based transcriptomic data uncovered 408,801 non-redundant transcripts with an average length of 1,685 bp. Of those, 189,406 transcripts were commonly identified by both methods, 169,149 transcripts with an average length of 619 bp were uniquely identified by Illumina HiSeq, and 51,246 transcripts with an average length of 2,535 bp were uniquely identified by PacBio Iso-Seq. Approximately 96 % of the final combined transcripts were mapped back to the Miscanthus genome, reflecting the high quality and coverage of our sequencing results. When comparing our data with genomes of four species of Andropogoneae, M. sinensis showed the closest relationship with sugarcane with up to 93 % mapping ratios, followed by sorghum with up to 80 % mapping ratios, indicating a high conservation of orthologs in these three genomes. Furthermore, 306,228 transcripts were successfully annotated against public databases including cell wall related genes and transcript factor families, thus providing many new insights into gene functions. The PacBio Iso-Seq data also helped identify 3,898 alternative splicing events and 2,963 annotated AS isoforms within 10 function categories. Conclusions Taken together, the present study provides a rich data set of full-length transcripts that greatly enriches our understanding of M. sinensis transcriptomic resources, thus facilitating further genetic improvement and molecular studies of the Miscanthus species. Supplementary Information The online version contains supplementary material available at 10.1186/s12864-021-07971-x.
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Affiliation(s)
- Yongli Wang
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China
| | - Xia Li
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China
| | - Congsheng Wang
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China
| | - Lu Gao
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China
| | - Yanfang Wu
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China
| | - Xingnan Ni
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China
| | - Jianzhong Sun
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China.
| | - Jianxiong Jiang
- Biofuels Institute, School of the Environment and Safety Engineering, Jiangsu University, 212013, Zhenjiang, Jiangsu, China.
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Decoding co-/post-transcriptional complexities of plant transcriptomes and epitranscriptome using next-generation sequencing technologies. Biochem Soc Trans 2021; 48:2399-2414. [PMID: 33196096 DOI: 10.1042/bst20190492] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2020] [Revised: 10/06/2020] [Accepted: 10/22/2020] [Indexed: 12/12/2022]
Abstract
Next-generation sequencing (NGS) technologies - Illumina RNA-seq, Pacific Biosciences isoform sequencing (PacBio Iso-seq), and Oxford Nanopore direct RNA sequencing (DRS) - have revealed the complexity of plant transcriptomes and their regulation at the co-/post-transcriptional level. Global analysis of mature mRNAs, transcripts from nuclear run-on assays, and nascent chromatin-bound mRNAs using short as well as full-length and single-molecule DRS reads have uncovered potential roles of different forms of RNA polymerase II during the transcription process, and the extent of co-transcriptional pre-mRNA splicing and polyadenylation. These tools have also allowed mapping of transcriptome-wide start sites in cap-containing RNAs, poly(A) site choice, poly(A) tail length, and RNA base modifications. The emerging theme from recent studies is that reprogramming of gene expression in response to developmental cues and stresses at the co-/post-transcriptional level likely plays a crucial role in eliciting appropriate responses for optimal growth and plant survival under adverse conditions. Although the mechanisms by which developmental cues and different stresses regulate co-/post-transcriptional splicing are largely unknown, a few recent studies indicate that the external cues target spliceosomal and splicing regulatory proteins to modulate alternative splicing. In this review, we provide an overview of recent discoveries on the dynamics and complexities of plant transcriptomes, mechanistic insights into splicing regulation, and discuss critical gaps in co-/post-transcriptional research that need to be addressed using diverse genomic and biochemical approaches.
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Zhang C, Ren H, Yao X, Wang K, Chang J. Full-length transcriptome analysis of pecan ( Carya illinoinensis) kernels. G3 GENES|GENOMES|GENETICS 2021; 11:6288450. [PMID: 34849807 PMCID: PMC8496322 DOI: 10.1093/g3journal/jkab182] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 02/25/2021] [Accepted: 05/18/2021] [Indexed: 11/12/2022]
Abstract
Abstract
Pecan is rich in bioactive components such as fatty acids (FAs) and flavonoids and is an important nut type worldwide. Therefore, the molecular mechanisms of phytochemical biosynthesis in pecan are a focus of research. Recently, a draft genome and several transcriptomes have been published. However, the full-length mRNA transcripts remain unclear, and the regulatory mechanisms behind the quality components biosynthesis and accumulation have not been fully investigated. In this study, single-molecule long-read sequencing technology was used to obtain full-length transcripts of pecan kernels. In total, 37,504 isoforms of 16,702 genes were mapped to the reference genome. The numbers of known isoforms, new isoforms, and novel isoforms were 9013 (24.03%), 26,080 (69.54%), and 2411 (6.51%), respectively. Over 80% of the transcripts (30,751, 81.99%) had functional annotations. A total of 15,465 alternative splicing (AS) events and 65,761 alternative polyadenylation events were detected; wherein, the retained intron was the predominant type (5652, 36.55%) of AS. Furthermore, 1894 long noncoding RNAs and 1643 transcription factors were predicted using bioinformatics methods. Finally, the structural genes associated with FA and flavonoid biosynthesis were characterized. A high frequency of AS accuracy (70.31%) was observed in FA synthesis-associated genes. This study provides a full-length transcriptome data set of pecan kernels, which will significantly enhance the understanding of the regulatory basis of phytochemical biosynthesis during pecan kernel maturation.
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Affiliation(s)
- Chengcai Zhang
- Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou 311400, China
| | - Huadong Ren
- Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou 311400, China
| | - Xiaohua Yao
- Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou 311400, China
| | - Kailiang Wang
- Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou 311400, China
| | - Jun Chang
- Research Institute of Subtropical Forestry, Chinese Academy of Forestry, Hangzhou 311400, China
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Yu H, Liu M, Yin M, Shan T, Peng H, Wang J, Chang X, Peng D, Zha L, Gui S. Transcriptome analysis identifies putative genes involved in triterpenoid biosynthesis in Platycodon grandiflorus. PLANTA 2021; 254:34. [PMID: 34291354 DOI: 10.1007/s00425-021-03677-2] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/17/2021] [Accepted: 06/30/2021] [Indexed: 05/25/2023]
Abstract
Comprehensive transcriptome analysis of different Platycodon grandiflorus tissues discovered genes related to triterpenoid saponin biosynthesis. Platycodon grandiflorus (Jacq.) A. DC. (P. grandiflorus), a traditional Chinese medicine, contains considerable triterpenoid saponins with broad pharmacological activities. Triterpenoid saponins are the major components of P. grandiflorus. Here, single-molecule real-time and next-generation sequencing technologies were combined to comprehensively analyse the transcriptome and identify genes involved in triterpenoid saponin biosynthesis in P. grandiflorus. We quantified four saponins in P. grandiflorus and found that their total content was highest in the roots and lowest in the stems and leaves. A total of 173,354 non-redundant transcripts were generated from the PacBio platform, and three full-length transcripts of β-amyrin synthase, the key synthase of β-amyrin, were identified. A total of 132,610 clean reads obtained from the DNBSEQ platform were utilised to explore key genes related to the triterpenoid saponin biosynthetic pathway in P. grandiflorus, and 96 differentially expressed genes were selected as candidates. The expression levels of these genes were verified by quantitative real-time PCR. Our reliable transcriptome data provide valuable information on the related biosynthesis pathway and may provide insights into the molecular mechanisms of triterpenoid saponin biosynthesis in P. grandiflorus.
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Affiliation(s)
- Hanwen Yu
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Mengli Liu
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Minzhen Yin
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Tingyu Shan
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Huasheng Peng
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
- Chinese Academy of Medical Sciences Research Unit (No. 2019RU057), National Resource Center for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, 100700, China
| | - Jutao Wang
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Xiangwei Chang
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Daiyin Peng
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China
| | - Liangping Zha
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China.
- Institute of Conservation and Development of Traditional Chinese Medicine Resources, Anhui Academy of Chinese Medicine, Hefei, 230012, China.
| | - Shuangying Gui
- College of Pharmacy, Anhui University of Chinese Medicine, Hefei, 230012, China.
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Peng Z, He Y, Parajuli S, You Q, Wang W, Bhattarai K, Palmateer AJ, Deng Z. Integration of early disease-resistance phenotyping, histological characterization, and transcriptome sequencing reveals insights into downy mildew resistance in impatiens. HORTICULTURE RESEARCH 2021; 8:108. [PMID: 33931631 PMCID: PMC8087834 DOI: 10.1038/s41438-021-00543-w] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/15/2020] [Revised: 03/11/2021] [Accepted: 03/22/2021] [Indexed: 05/11/2023]
Abstract
Downy mildew (DM), caused by obligate parasitic oomycetes, is a destructive disease for a wide range of crops worldwide. Recent outbreaks of impatiens downy mildew (IDM) in many countries have caused huge economic losses. A system to reveal plant-pathogen interactions in the early stage of infection and quickly assess resistance/susceptibility of plants to DM is desired. In this study, we established an early and rapid system to achieve these goals using impatiens as a model. Thirty-two cultivars of Impatiens walleriana and I. hawkeri were evaluated for their responses to IDM at cotyledon, first/second pair of true leaf, and mature plant stages. All I. walleriana cultivars were highly susceptible to IDM. While all I. hawkeri cultivars were resistant to IDM starting at the first true leaf stage, many (14/16) were susceptible to IDM at the cotyledon stage. Two cultivars showed resistance even at the cotyledon stage. Histological characterization showed that the resistance mechanism of the I. hawkeri cultivars resembles that in grapevine and type II resistance in sunflower. By integrating full-length transcriptome sequencing (Iso-Seq) and RNA-Seq, we constructed the first reference transcriptome for Impatiens comprised of 48,758 sequences with an N50 length of 2060 bp. Comparative transcriptome and qRT-PCR analyses revealed strong candidate genes for IDM resistance, including three resistance genes orthologous to the sunflower gene RGC203, a potential candidate associated with DM resistance. Our approach of integrating early disease-resistance phenotyping, histological characterization, and transcriptome analysis lay a solid foundation to improve DM resistance in impatiens and may provide a model for other crops.
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Affiliation(s)
- Ze Peng
- University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA
- State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, South China Agricultural University, 510642, Guangzhou, China
| | - Yanhong He
- Visiting scholar at University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA
- Key Laboratory of Horticultural Plant Biology, Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, 430070, Wuhan, Hubei, China
| | - Saroj Parajuli
- University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA
| | - Qian You
- University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA
| | - Weining Wang
- University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA
| | - Krishna Bhattarai
- University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA
| | - Aaron J Palmateer
- University of Florida, IFAS, Department of Plant Pathology, Tropical Research and Education Center, 18905 S.W. 280th Street, Homestead, FL, 33031, USA
- Bayer Environmental Science US, 5000 Centregreen Way, Cary, NC, 27513, USA
| | - Zhanao Deng
- University of Florida, IFAS, Department of Environmental Horticulture, Gulf Coast Research and Education Center, 14625 County Road 672, Wimauma, FL, 33598, USA.
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Ramberg S, Høyheim B, Østbye TKK, Andreassen R. A de novo Full-Length mRNA Transcriptome Generated From Hybrid-Corrected PacBio Long-Reads Improves the Transcript Annotation and Identifies Thousands of Novel Splice Variants in Atlantic Salmon. Front Genet 2021; 12:656334. [PMID: 33986770 PMCID: PMC8110904 DOI: 10.3389/fgene.2021.656334] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2021] [Accepted: 04/01/2021] [Indexed: 12/18/2022] Open
Abstract
Atlantic salmon (Salmo salar) is a major species produced in world aquaculture and an important vertebrate model organism for studying the process of rediploidization following whole genome duplication events (Ss4R, 80 mya). The current Salmo salar transcriptome is largely generated from genome sequence based in silico predictions supported by ESTs and short-read sequencing data. However, recent progress in long-read sequencing technologies now allows for full-length transcript sequencing from single RNA-molecules. This study provides a de novo full-length mRNA transcriptome from liver, head-kidney and gill materials. A pipeline was developed based on Iso-seq sequencing of long-reads on the PacBio platform (HQ reads) followed by error-correction of the HQ reads by short-reads from the Illumina platform. The pipeline successfully processed more than 1.5 million long-reads and more than 900 million short-reads into error-corrected HQ reads. A surprisingly high percentage (32%) represented expressed interspersed repeats, while the remaining were processed into 71 461 full-length mRNAs from 23 071 loci. Each transcript was supported by several single-molecule long-read sequences and at least three short-reads, assuring a high sequence accuracy. On average, each gene was represented by three isoforms. Comparisons to the current Atlantic salmon transcripts in the RefSeq database showed that the long-read transcriptome validated 25% of all known transcripts, while the remaining full-length transcripts were novel isoforms, but few were transcripts from novel genes. A comparison to the current genome assembly indicates that the long-read transcriptome may aid in improving transcript annotation as well as provide long-read linkage information useful for improving the genome assembly. More than 80% of transcripts were assigned GO terms and thousands of transcripts were from genes or splice-variants expressed in an organ-specific manner demonstrating that hybrid error-corrected long-read transcriptomes may be applied to study genes and splice-variants expressed in certain organs or conditions (e.g., challenge materials). In conclusion, this is the single largest contribution of full-length mRNAs in Atlantic salmon. The results will be of great value to salmon genomics research, and the pipeline outlined may be applied to generate additional de novo transcriptomes in Atlantic Salmon or applied for similar projects in other species.
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Affiliation(s)
- Sigmund Ramberg
- Department of Life Sciences and Health, Faculty of Health Sciences, OsloMet - Oslo Metropolitan University, Oslo, Norway
| | - Bjørn Høyheim
- Department of Preclinical Sciences and Pathology, Faculty of Veterinary Medicine, Norwegian University of Life Sciences, Ås, Norway
| | | | - Rune Andreassen
- Department of Life Sciences and Health, Faculty of Health Sciences, OsloMet - Oslo Metropolitan University, Oslo, Norway
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Zhao C, He L, Xia H, Zhou X, Geng Y, Hou L, Li P, Li G, Zhao S, Ma C, Tang R, Pandey MK, Varshney RK, Wang X. De novo full length transcriptome analysis of Arachis glabrata provides insights into gene expression dynamics in response to biotic and abiotic stresses. Genomics 2021; 113:1579-1588. [PMID: 33819563 DOI: 10.1016/j.ygeno.2021.03.030] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2020] [Revised: 03/21/2021] [Accepted: 03/29/2021] [Indexed: 11/26/2022]
Abstract
The perennial ornamental peanut Arachis glabrata represents one of the most adaptable wild Arachis species. This study used PacBio combined with BGISEQ-500 RNA-seq technology to study the transcriptome and gene expression dynamics of A. glabrata. Of the total 109,747 unique transcripts obtained, >90,566 transcripts showed significant homology to known proteins and contained the complete coding sequence (CDS). RNA-seq revealed that 1229, 1039, 1671, 3923, 1521 and 1799 transcripts expressed specifically in the root, stem, leaf, flower, peg and pod, respectively. We also identified thousands of differentially expressed transcripts in response to drought, salt, cold and leaf spot disease. Furthermore, we identified 30 polyphenol oxidase encoding genes associated with the quality of forage, making A. glabrata suitable as a forage crop. Our findings presented the first transcriptome study of A. glabrata which will facilitate genetic and genomics studies and lays the groundwork for a deeper understanding of the A. glabrata genome.
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Affiliation(s)
- Chuanzhi Zhao
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China; College of Life Sciences, Shandong Normal University, Jinan 250014, People's Republic of China.
| | - Liangqiong He
- Cash Crop Research Institute, Guangxi Academy of Agricultural Sciences, 530007 Nanning, People's Republic of China
| | - Han Xia
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China; College of Life Sciences, Shandong Normal University, Jinan 250014, People's Republic of China
| | - Ximeng Zhou
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China; College of Life Sciences, Shandong Normal University, Jinan 250014, People's Republic of China
| | - Yun Geng
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China
| | - Lei Hou
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China
| | - Pengcheng Li
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China; College of Life Sciences, Shandong Normal University, Jinan 250014, People's Republic of China
| | - Guanghui Li
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China
| | - Shuzhen Zhao
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China
| | - Changle Ma
- College of Life Sciences, Shandong Normal University, Jinan 250014, People's Republic of China
| | - Ronghua Tang
- Cash Crop Research Institute, Guangxi Academy of Agricultural Sciences, 530007 Nanning, People's Republic of China
| | - Manish K Pandey
- Center of Excellence in Genomics & Systems Biology (CEGSB), International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Hyderabad 502324, India
| | - Rajeev K Varshney
- Center of Excellence in Genomics & Systems Biology (CEGSB), International Crops Research Institute for the Semi-Arid Tropics (ICRISAT), Hyderabad 502324, India; State Agricultural Biotechnology Centre, Centre for Crop and Food Innovation, Food Futures Institute, Murdoch University, Murdoch, Western Australia, Australia
| | - Xingjun Wang
- Biotechnology Research Center, Shandong Academy of Agricultural Sciences, Shandong Provincial Key Laboratory of Crop Genetic Improvement, Ecology and Physiology, Jinan 250100, People's Republic of China; College of Life Sciences, Shandong Normal University, Jinan 250014, People's Republic of China.
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Ni L, Wang Z, Guo J, Pei X, Liu L, Li H, Yuan H, Gu C. Full-Length Transcriptome Sequencing and Comparative Transcriptome Analysis to Evaluate Drought and Salt Stress in Iris lactea var. chinensis. Genes (Basel) 2021; 12:434. [PMID: 33803672 PMCID: PMC8002972 DOI: 10.3390/genes12030434] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2021] [Revised: 03/15/2021] [Accepted: 03/16/2021] [Indexed: 12/12/2022] Open
Abstract
Iris lactea var. chinensis (I. lactea var. chinensis) is a perennial herb halophyte with salt and drought tolerance. In this study, full-length transcripts of I. lactea var. chinensis were sequenced using the PacBio RSII sequencing platform. Moreover, the transcriptome was investigated under NaCl or polyethylene glycol (PEG) stress. Approximately 30.89 G subreads were generated and 31,195 unigenes were obtained by clustering the same isoforms by the PacBio RSII platform. A total of 15,466 differentially expressed genes (DEGs) were obtained under the two stresses using the Illumina platform. Among them, 9266 and 8390 DEGs were obtained under high concentrations of NaCl and PEG, respectively. In total, 3897 DEGs with the same expression pattern under the two stresses were obtained. The transcriptome expression profiles of I. lactea var. chinensis under NaCl or PEG stress obtained in this study may provide a resource for the same and different response mechanisms against different types of abiotic stress. Furthermore, the stress-related genes found in this study can provide data for future molecular breeding.
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Affiliation(s)
- Longjie Ni
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
- College of Forest Sciences, Nanjing Forestry University, Nanjing 210037, China;
| | - Zhiquan Wang
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
| | - Jinbo Guo
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
| | - Xiaoxiao Pei
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
| | - Liangqin Liu
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
| | - Huogen Li
- College of Forest Sciences, Nanjing Forestry University, Nanjing 210037, China;
| | - Haiyan Yuan
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
| | - Chunsun Gu
- Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China; (L.N.); (Z.W.); (J.G.); (X.P.); (L.L.); (H.Y.)
- College of Forest Sciences, Nanjing Forestry University, Nanjing 210037, China;
- Jiangsu Provincial Platform for Conservation and Utilization of Agricultural Germplasm, Nanjing 210014, China
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