Germline mutation analysis of MLH1 and MSH2 in Malaysian Lynch syndrome patients

Table 1 List of primer sequences used for polymerase chain reaction amplification of the exons of MLH1 gene and their respective sizes of polymerase chain reaction products

MLH1 gene	Primer sequence (5’-3’)	Size (bp)	Ta (°C)
Exon 1	5’ cagagttgagaaatttgactgg 3’	339	50
	5’ taagtcgtagcccttaagtgag 3’
Exon 2	5’ tgtatgagcctgtaagacaaag 3’	315	50
	5’ tagtttccagaacagagaaagg 3’
Exon 3	5’ gggaattcaaagagatttgg 3’	373	50
	5’ gaggtctctgcaggtaaaatag 3’
Exon 4	5’ ctcatcgttgccacatatta 3’	319	50
	5’ gtactcaagatctctgccaaaa 3’
Exon 5	5’ tccccttgggattagtatctat 3’	306	50
	5’ cccacaaaagccaatagtcat 3’
Exon 6	5’ cactatcttaagacctcgcttt 3’	325	50
	5’ gctaaaccttgaccagaaacta 3’
Exon 7	5’ agaggagtctgtgttttgattc 3’	367	50
	5’ atcataaccttatctccaccag 3’
Exon 8	5’ atagtttgctggtggagataag 3’	306	50
	5’ agcctgtgtatttgactaaagc 3’
Exon 9	5’ agttagtttatgggaaggaacc 3’	438	50
	5’ aagaaccagactccaacagtc 3’
Exon 10	5’ atagtaagatagtgggctggaa 3’	432	62
	5’ tacctgtaagaagggacagaac 3’
Exon 11	5’ tatttgaacacactcagactcg 3’	476	62
	5’ taggaacaacagcacaatacc 3’
Exon 12	5’ atacagactttgctaccaggac 3’	475	50
	5’ agagagaagatgcaagtgattc 3’
Exon 13	5’ tgtcagataagcagtctaccag 3’	468	50
	5’ ttagtaaaggaagaggagcttg 3’
Exon 14	5’ cctcatttgtgttcgttttc 3’	411	50
	5’ gtcgaacttggatttgaaac 3’
Exon 15	5’ catgtttcagggattacttctc 3’	356	50
	5’ ggctaccaaatgactattttgc 3’
Exon 16	5’ gtatatgggggatgagtgttac 3’	319	50
	5’ cagaagtataagaatggctgtc 3’
Exon 17	5’ gtgctgtttgtcaggatgaa 3’	430	50
	5’ tgaatattagtggggatgagag 3’
Exon 18	5’ gtctgtgatctccgtttagaat 3’	358	50
	5’ ctacatcagcagatggaagtaa 3’
Exon 19	5’ caggacaccagtgtatgttg 3’	497	50
	5’ ctgtgacatgttcaagacctc 3’

bp: Base pair; Ta: Annealing temperature.

Table 2 List of primer sequences used for polymerase chain reaction amplification of the exons of MSH2 gene and their respective sizes of polymerase chain reaction products

MSH2 gene	Primer sequence (5’-3’)	Size (bp)	Ta (°C)
Exon 1	5’ gtctgcttatgattggttgc 3’	632	58
	5’ actccgtgatcacaagttca 3’
Exon 2	5’ gtggaactataagtccagtaagc 3’	457	50
	5’ cagccaaactgcaacttttt 3’
Exon 3	5’ agaatcgattgaacccttga 3’	554	62
	5’ gtcaattaaagagcctttcc 3’
Exon 4	5’ acatcatatcagtgtcttgcac 3’	302	50
	5’ taatccatgtacctgattctcc 3’
Exon 5	5’ gaaggaacaccaaggaaaat 3’	273	50
	5’ ctgaaaaaggttaagggctctg 3’
Exon 6	5’ ggtcttaggaagaggaactttt 3’	337	50
	5’ gtataatcatgtgggtaactgc 3’
Exon 7	5’ ttgagacttacgtgcttagttg 3’	391	50
	5’ tctgaatgtgtcctaagagtga 3’
Exon 8	5’ aactttggagacctactgtact 3’	353	50
	5’ ccacaaaggtgctacaattag 3’
Exon 9	5’ tacatcatcagcactgtaactg 3’	494	53
	5’ ttgacagagatgtgaagtcatc 3’
Exon 10	5’ cagctttcaagtcagaaact 3’	511	53
	5’ aagaaagcttgactcttacctg 3’
Exon 11	5’ tcacgtagtacacattgcttct 3’	365	61
	5’ tatgtgcaagagtaacttccag 3’
Exon 12	5’ caaatggggggattaaatgt 3’	377	50
	5’ aaaacgttacccccacaaag 3’
Exon 13	5’ gcagtaactctgtccacatctt 3’	432	62
	5’ cttctcacaggacagagacata 3’
Exon 14	5’ atgtttgtggcatatccttc 3’	499	50
	5’ cagactgtgaattaaggggtaa 3’
Exon 15	5’ ctaatgacaaggtgagaaggat 3’	338	50
	5’ ccttcatcttagtgtcctgttt 3’
Exon 16A	5’ ggaactagacagtgcacacata 3’	428	62
	5’ ctgggatttttcacgtagtaac 3’
Exon 16B	5’ attattcaggagttcctgtcc 3’	500	50
	5’ ctctgagctatttgtttcttcc 3’

bp: Base pair; Ta: Annealing temperature.

Table 3 Germline mutations of MSH2 gene detected in 4/34 Lynch syndrome patients

Gene	Patients ID	Age	Exon/intron region	Nucleotide change	Consequence
MSH2	LS8	50	Exon 12	c.2005G > C	Missense mutation (G669R)
	LS20	43	Exon 12	c.2005G > C	Missense mutation (G669R)
	LS37	41	Intron 12	c.2006-6T > C	Splice-site mutation
	LS41	49	Exon 1	c.142G > T	Nonsense mutation (E48X)