Screening of Minor Mutation of hMLH1 and hMSH2 Based on Denatured HPLC

Objective:To establish a method based on denatured high performance liquid chromatography(DHPLC)and quickly detect minor mutation of mismatch repair gene hMLH1 and hMSH2.Methods:The primers to amplify all exons of hMLH1 and hMSH2 were designed by ourselves.Polymerase chain reaction(PCR)and DHPLC were deployed to screen minor mutation of hMLH1 and hMSH2.The results were compared with previous results of DNA sequencing to screen mutation of these genes.Results:All primers of hMLH1 and hMSH2 amplified corresponding exons and splicing regions of these genes very well.DHPLC detected all known mutations of these genes and sensitivity and specificity of screening mutation and nonmutation were all 100 %.Two temperatures were used to screen the mutation in some exons and splicing regions of hMLH1 and hMSH2 such as amplicon 12A of hMLH1 and amplicon 2,3,5 and 7 ofhMSH2.In contrast with DNA sequencing,DHPLC was quicker,highly efficient,with low work intensity,low-cost,low personal error,high sensitivity and specificity.Conclusion:The platform based on DHPLC can efficiently screen minor mutation of hMLH1 and hMSH2,and has high cost efficiency.