Parentally imprinted allele (PIA) typing in the differentially methylated region upstream of the human H19 gene.
Parentally imprinted allele (PIA) typing in the differentially methylated region upstream of the human H19 gene.
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DOI：
10.1016/j.fsigen.2008.03.008
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发表时间：
2008-09
期刊：
影响因子：
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通讯作者：
Daixin Huang;Xiao-jun Lin;Hui Chen;Qingen Yang;Ya Jie;Xian-dun Zhai;Hui Yin
中科院分区：
文献类型：
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作者：
Daixin Huang;Xiao-jun Lin;Hui Chen;Qingen Yang;Ya Jie;Xian-dun Zhai;Hui Yin
H19基因是位于染色体11p15.5上的父系印记基因。在本研究中，根据GenBank序列（登录号AF125183），研究了H19FR1和H19FR2单倍型多态性，分别包括H19基因上游的四个和三个SNP。通过PCR和随后的变性梯度凝胶电泳（DGGE），在中国汉族人群中检测到H19FR1的5种单倍型和9种基因型。辨别力 (Dp)、多态性信息含量 (PIC) 和亲子排除概率 (PE) 估计分别为 0.803、0.58 和 0.322。对于H19FR2，观察到2个单倍型和3个基因型，Dp、PIC和PE分别为0.626、0.37和0.162。测序结果显示，在中国汉族人群的H19FR1中仅检测到已报道的4个SNP中的2个，a7342g和g7547a，并发现了2个新的SNP，g7351c和a7357g。在 H19FR2 区域，仅检测到三个报告的 SNP 之一：a8097g。根据基因组 DNA 的甲基化状态，使用两种类型的酶（甲基化敏感限制性酶 (msRE) HpaII 或 HhaI 和 McrBC）对 H19FR 亲本等位基因进行选择性检测。经 HpaII 或 HhaI 消化的基因组 DNA 显示出一条源自父本等位基因的单条带，这是母本等位基因上未甲基化识别位点裂解的结果。相反，使用可以消化甲基化父本序列的McrBC，导致专门扩增母本等位基因。这种亲本印记等位基因（PIA）分型方法可能是区分等位基因亲本起源的有用技术之一。
The H19 gene is a paternally imprinted gene located on chromosome 11p15.5. In this study, the H19FR1 and H19FR2 haplotype polymorphisms including four and three SNPs, respectively, upstream of the H19 gene according to the GenBank sequence (accession no. AF125183) were investigated. Five haplotypes and nine genotypes were detected for H19FR1 in the Chinese Han population by means of PCR and subsequent denaturing gradient gel electrophoresis (DGGE). The power of discrimination (Dp), polymorphism information content (PIC) and probability of paternity exclusion (PE) were estimated to be 0.803, 0.58 and 0.322, respectively. For the H19FR2, two haplotypes and three genotyes were observed, and the Dp, PIC and PE were 0.626, 0.37 and 0.162, respectively. Sequencing results showed that only two of the four reported SNPs, a7342g and g7547a, were detected in H19FR1 in the Chinese Han population, and two new SNPs, g7351c and a7357g, were found. In the H19FR2 region, only one of the three reported SNPs, a8097g, was detected. Based on the methylation status of the genomic DNA, selective detection of the parental alleles for H19FRs was examined by using two types of enzymes, the methylation-sensitive restriction enzyme (msRE) HpaII or HhaI and McrBC. Genomic DNA digested by either HpaII or HhaI, revealed a single band derived from the paternal allele, as a result of cleavage of unmethylated recognition sites on the maternal allele. On the contrary, the use of McrBC, which can digest a methylated paternal sequence, resulted in exclusively amplifying the maternal allele. This parentally imprinted allele (PIA) typing method could be one of the useful techniques for discriminating the parental origin of alleles.