Three novel missense germline mutations in different exons of MSH6 gene in Chinese hereditary non-polyposis colorectal cancer families

Table 1 Primer sequences and PCR condition of different exons of MSH6 gene

EN	Primer sequence (5’-3’)	Size(bp)	AT(°C)	CN(C)
Exon1	M13F-AGCTCCGTCCGACAGAAC	381	68	38
	M13R-CTGTGCGAGCCTCCCCT
Exon2	M13F-TGCCAGAAGACTTGGAATTG	325	63	32
	M13R-CAAACACACACACATGGCAG
Exon3	M13F-GATGGGGTTTGCTATGTTGC	341	67	36
	M13R-TACACCCTCCCCCTTTCTTC
Exon4.1	M13F-GGCTGCACGGGTACCATTAT	390	60	34
	M13R-CATTCTCTTCCGCTTTCGAG
Exon4.2	M13F-GCCAGACACTAAGGAGGAAGG	386	59	35
	M13R-TAGATGCATCAAAATCGGGG
Exon4.3	M13F-TGGCTTAAGGAGGAAAAGAGA	378	60	34
	M13R-TCTACATCGTGCCTCCATCA
Exon4.4	M13F-TTCTGGCTTTCCTGAAATTG	374	60	34
	M13R-TAAATCTCGAACAATGGCGA
Exon4.5	M13F-TCTGGCCATACTCGTGCATA	329	60	34
	M13R-AGCACCTGGGGTAACATCAC
Exon4.6	M13F-TCAGGAAGGTCTGATACCCG	353	62	33
	M13R-GCACCATTCGTTGATAGGCT
Exon4.7	M13F-AAGTGAATTGGCCCTCTCTG	483	62	30
	M13R-TGGTTCTGACTCTTCAGGGG
Exon4.8	M13F-TTTTGGTAAGCGGCTCCTAA	465	62	35
	M13R-TTTCGAGCCTTTTCATGGTC
Exon4.9	M13F-TTTCTGCTCTGGAAGGATTC	440	62	30
	M13R-TCGTTTACAGCCCTTCTTGG
Exon4.10	M13F-TGAACAGAGCCTCCTGGAAT	390	62	32
	M13R-CAGCTGGCAAACAGCACTAC
Exon5	M13F-CTGATAAAACCCCCAAACGA	403	62	30
	M13R-CTGTGTTTGGAAAATGATCACC
Exon6	M13F-CCAGTCATAAAAGACCTTTTCC	192	56	34
	M13R-GACTGAATGAGAACTTAAGTGGG
Exon7	M13F-AAGGTGAAAGTACATTT	119	61	33
	M13R-TTCAAATGAGAAGTTTAATG
Exon8/9	M13F-CCTTTGAGTTACTTCCTT	573	61	32
	M13R-TCATAGTGCATCATCCCTTCC
Exon10	M13F-GGAAGGGATGATGCACTATG	254	61	35
	M13R-AAGAAAATGGAAAAATGGTCA

Abbreviation: EN, Exon name. AT, Anneal temperature. CN, Cycle number.The sequence of M13F was 5’-GTAAAACGACGGCCAGT-3’. The sequence of M13R was 5’-AACAGCTATGACCATG-3’.

Table 2 Germline mutations of MSH6 gene in 4 probands of HNPCC families

FamilyNo	Exon	Position ofmutation	Base change	Result	Mutation type
H3	4.3	Codon 468	c.1403G>A	Arg>His	Missense mutation
H14	6	Codon 1163	c.3488A>T	Glu>Val	New SNPs
H40	4.6	Codon 666	c.1996T>C	Ser>Pro	Missense mutation
H61	9	Codon 1284	c.3851C>T	Thr>Met	Missense mutation

Table 3 Clinical characteristics of 4 mutational probands of HNPCC families

No	Sex	Age(yr)	Criteria	Site of cancer	Age at diagnosis of first CRC (yr)	Metachronous tumor
H3	M	50	AC	Sigmoid	46	VA
H14	F	26	BG	Rectum	26	NMT
H40	F	39	BG	Rectum	39	NMT
H61	F	51	BG	Descending	33	EC

Abbreviation: AC, Amsterdam criteria. JC, Japanese criteria. BG, Bethesda guidelines. CRC, Colorectal cancer. VA, Villiform adenoma. NMT: No Metachronous tumor. EC, Endometrial cancer.