基于二代测序平台的微单倍型位点的分型研究

Genotyping of microhaplotypes based on the second generation sequencing platform

目的建立基于二代测序平台的微单倍型分型方案,调查其在中国南方汉族人群中的多态性,探讨其法医学应用价值。方法以20个微单倍型位点为研究对象,利用多重PCR构建靶向扩增子文库,以二代测序为检测手段,采用生物信息学方法构建个体微单倍型,最后进行多态性分析。结果总体测序量为2.12G,覆盖所有片段,平均覆盖深度为3353×。微单倍型内含72个SNP位点的观测杂合度范围为0.109~0.587,平均值为0.364;而20个微单倍型位点本身的观测杂合度范围为0.198~0.837,平均值为0.726,均高于SNP位点。20个微单倍型位点中仅一个位点未通过Hardy-Weinberg平衡检验,剩余19个位点的有效等位基因数值为3.119~7.170,多态信息含量为0.620~0.845,平均多态信息含量为0.711。19个有效微单倍型位点的匹配概率为0.044~0.181,累积匹配概率为3.132×10^(-19)。结论本次研究证实了19个微单倍型的参数效能与常用15个STR基因座相当,可独立应用于法医个体识别并具潜在混合物分析价值,该结果为法医遗传学实践提供更多选择。

Objective To investigate the genetic polymorphism of 20 microhaplotypes in Chinese South Han population and assess their value in forensic medicine based on the second generation sequencing.Methods 20 microhaplotype loci were selected as the research objects,and the targeted amplified fragment library was constructed by multiplex PCR.Amplicon and The individual microhaplotypes were constructed by Bioinformatics Method with the second generation sequencing as detection method,and the polymorphism was analyzed.Results A total of 2.12 G original data were obtained,covering all the fragments with average depth of 3353×.The observed heterozygosity of 20 microhaplotypes ranged from 0.198 to 0.837 with an average of 0.726,while that of their SNP ranged from 0.109 to 0.587 with an average of 0.364.Only 1 microhaplotype cannot pass the Hardy-Weinberg equilibrium test.The effective number of alleles values of the remaining 19 microhaplotypes ranged from 3.119 to 7.170 and the polymorphic information content ranged from 0.620 to 0.845 with an average of 0.711.The matching probability of the 19 microhaplotypes ranged from 0.044 to 0.181 and the cumulative matching probability value was 3.132×10^(-19).Conclusion This study confirmed that the performance of 19 microhaplotype systems for forensic parameters is equivalent to the commonly used 15 STR loci,which can be independently applied to forensic individual identification and the mixed DNA genotyping.