用分子克隆技术构建D12S375基因座等位基因分型标准物及汉、回、维族群体遗传学研究

Study on the construction of standard D12S375 allelic ladder and its genetic polymorphism in Chinese Han, Hui and Wei populations.

采用分子克隆技术大量制备STR基因座等位基因分型标准物 ,解决长期困扰STR分型的准确性和标准化问题。PCR扩增出D12S375基因座的 7个等位基因片段 ,将其插入pUC重组质粒中 ,经DNA测序分析证实插入片段的结构及大小 ,按重复单位的重复次数对插入的等位基因片段进行命名 ,最后经转染、扩增及再鉴定后制备出标准D12S375基因座等位基因分型标准物。应用此分型标准物 ,调查D12S375基因座在成都汉族 ,甘肃回族及新疆维族群体中的基因型分布频率 ,评估其在法医学实践中的应用价值。

s:To resolve the problem of the accuracy and standardization of STR PCR typing in forensic science practice,we have designed a new method to produce standard D12S375 allelic ladder.Seven different PCR amplified D12S375 allelic fragments were isolated from the gel,eluted into the distilled water and reamplified by PCR.The purified allelic fragments were then blunt end subcloned individually into the pUC plasmid vectors and transfected into competent E.coli DH5α TM cells.The sequencing results confirmed that the size and the constructure of the inserts were correct.The recombinant plasmids DNA with 7 inserts were then used as templates for reamplification to generate D12S375 standard ladder, with which the genetic polymorphisms of D12S375 locus in Chinese Han population in Chengdu,Hui population in Gansu and Wei population in Xinjiang were studied.