应用Ion Torrent PGM^TM平台进行人mtDNA全基因组测序

目的应用Ion Torrent PGM^TM平台对人线粒体基因组全序列进行分析检测。方法采集39名辽宁汉族无关个体以及4个母系家系的14名相关个体样本,应用SequalPrep^TM Long PCR试剂盒进行扩增,应用Ion Shear^TM Plus Reagents试剂盒和Ion Plus Fragment Library试剂盒等构建文库,并在Ion Torrent PGM平台上进行线粒体基因组全序列测序。结果在39名无关个体共观察到39种单倍型,在396个位置观察到了397种碱基变异。无关个体中出现的变异位点数目为25~53个,平均每个个体出现36.2个碱基变异。4个母系家系中每个家系成员间具有完全相同的mtDNA单倍型,严格遵守母系遗传。结论采用本研究建立的人线粒体基因组全序列的测序检验法,可显著提高mtDNA的个体识别能力,在法庭科学领域中有较好的应用价值。

The application of whole human mitochondriai genomes sequencing using Ion Torrent PGM^TM platform

Objective Based on the next generation sequencing （NGS） Ion Torrent Personal Genomic Machine^TM（PGM） platform, we demonstrated a new sequencing strategy for entire human mitochondrial genomes （mtGenomes）. Methods Samples were collected from 39 unrelated Liaoning Han individuals and 4 maternal families. Full mtGenomes were amplified by SequalPrepTM Long PCR kit. Libraries were constructed with Ion ShearTM Plus Reagents kit and Ion Plus Fragment Library kit. NGS was performed using PGM. All PGM sequences were analyzed with the Ion Torrent Software Suite. Results Among 39 unrelated individuals, a total of 39 different haplotypes were observed and 397 variable sites were found. Mutation sites varied from 25 to 53 with the 36.2 averaged. In the 4 maternal families, the mitochondrial DNA （mtDNA） haplotype was identical from the same maternal individuals, which was in strict conformity with maternal inheritance mode. Conclusion This sequencing strategy allows for the greater power discrimination of mtDNA typing and provides a theoretical basis for further applications in forensic science.