A study on SNP polymorphisms in the hypermethylated region upstream of the human H19 gene / 中国法医学杂志

ABSTRACT

Objective To establish a simple and high-performance analytical technique for detecting DNA methylation markers and SNPs simultaneously,and obtain the population genetics data of some SNPs in the hypermethylated region upstream of the human H19 gene.Methods The haplotypes of H19FR1 and H19FR2 which located in the promoter region upstream of the human H19 gene were investigated from 232 unrelated Chinese individuals living in Wuhan by means of PCR and subsequent denaturing gradient gel electrophoresis(DGGE).Based on the methylation status of the genomic DNA,selective detection of the parental alleles for H19FRs was examined by using the methylation-sensitive restriction enzyme(msRE) Hpa II or Hha I.Results Five haplotypes and nine phenotypes were observed for H19FR1 in Chinese Han population in Wuhan,and the power of discrimination(DP),polymorphism information content(PIC) and probability of paternity exclusion(PE) were 0.803,0.58 and 0.322 respectively.For the H19FR2,two haplotypes and three phenotyes were detected,and the DP,PIC and PE were 0.626,0.37 and 0.162 respectively.Sequencing results showed that there were 3 SNPs,a7342g,a7357g and g7547a,and one g7351c point mutation in H19FR1.In the H19FR2,there was only one SNP,a8097g.The msRE,HpaⅡ or HhaⅠ,could digest the maternal allele,and only a single band derived from the paternal allele was detected by post-digestion PCR-DGGE(PDP-DGGE) technique.Conclusion PDP-DGGE is a simple,sensitive and effective technique for analyzing DNA methylation status and SNPs simultaneously,and can be used for discriminating the parental origin of alleles.