Single-tube-genotyping of gastric cancer related SNPs by directly using whole blood and paper-dried blood as starting materials

Figure 1 Schematic of a single-tube-genotyping method using tetra-PCR amplification from blood and extracted DNA.

Figure 2 Direct PCR amplification from blood samples with the anticoagulant of dipotassium EDTA (1, 4); Sodium citrate (2, 5); and Sodium heparinate (3, 6).

Figure 3 Direct PCR with two sets of primers (P1-1 and P1-2; P2-1 and P2-2) using 0. 5 μL (a1, b1), 1 μL (a2, b2), 2 μL (a3, b3) and 4 μL (a4, b4) of EDTA-blood dried on filter paper as starting materials. PCR with 1 μL of fresh blood without an anticoagulant was performed in parallel for the comparison (a5, b5). a1-a5: 379-bp fragment; b1-b5: 539-bp fragment.

Figure 4 Results of genotyping of IL-1B-31 using whole blood as the starting PCR material.

Figure 5 Results of genotyping of IL-1B-31 using paper-dried blood as starting PCR material.

Figure 6 The optimized experiment results of polymorphism IL-1B-31 by changing the concentration ratios of two allele-specific primers from 1:1 to 8:1 (S1-1: S1-2) to get relatively equal band-intensities. A: PCR using whole blood as starting PCR materials; B: PCR using paper-dried blood as starting PCR materials.

Figure 7 Results of genotyping of IL-1B-31 using extracted genomic DNA as starting PCR materials.

Figure 8 Results of genotyping of IL-1B-511 using whole blood as starting PCR materials.

Figure 9 Electrophoresis results of three typical genotypes were obtained, heterozygote (C/T) and both homozygotes (T/T and C/C) of IL-1B-31, by the microchip (A) and Sanger’s sequencing method (B) respectively. The fragment of 379 bp was from two outer primers (P1-1 and P1-2).

Figure 10 Electrophoresis results of three typical genotypes were obtained, heterozygote (C/T) and both homozygotes (T/T and C/C) of IL-1B-511, by the microchip (A) and Sanger’s sequencing method (B) respectively. The fragment of 539 bp was from two outer primers (P2-1 and P2-2).