基于微流控芯片的InDel快速族群推断体系研究

Study of The InDel Ancestry Inference Microfluidic Chip Amplification System

目的QuickTargSeq全集成法医DNA现场快速检测系统是国内首台自主研制的现场快检仪,可应用于InDel族群推断检测,2 h左右完成“样本进-结果出”的快速自动化InDel分型。本文对InDel族群推断微流控芯片检测体系的性能进行评估,以期为实践应用提供参考。方法使用InDel族群推断微流控芯片检测体系,对体系的灵敏度、干扰物耐受性、成功率、分型准确率、精确性、准确性、峰平衡性及检材适应性进行验证评估,同时对测试样本的族群来源进行推断。结果138份样本的全集成检测成功率为95.65%,分型准确率为98.85%;DNA模板量≥5 ng时,可获得完整InDel分型,口腔拭子样本最佳采集次数为口腔内壁左右两侧各刮擦8次,血卡样本最佳检测方式为6片(Φ=2 mm);所有基因座的平均杂合子峰高比值为0.86;10次运行的等位基因分型标准物(allelic ladder)片段大小标准差均在0.3 bp以内,测试样本等位基因和相应的等位基因分型标准物之间的片段准确性均在0.5 bp以内。结论该体系可实现对口腔拭子、血卡、唾液卡及烟蒂样本的准确分型,能够准确推断样本的族群来源。

Objective Quick TarSeq integrated system developed by our groups for on-field fast analysis is a fully automated and integrated system,which can be applied to the InDel ancestry inference,and complete the“sample-in result-out”rapid automatic InDel typing in about 2 h.In this paper,the performance of the InDel ancestry inference microfluidic chip amplification system was studied,hoping to provide a reference for practical application.Methods The system was validated by sensitivity,inhibitor tests,success rate,concordance tests,sizing precision and accuracy,peak height balance,case sample tests,and the accuracy of infering ancestral origins.Results The microfluidic chip amplification system was concordant with traditional methods,with 95.65%success rates and 98.85%concordance rates for 138 testing samples.The complete InDel profiles can be obtained from more than 5 ng of template DNA input.The best collection of buccal swab was the donor who performed 8 swipes inside of cheek.The best collection of dried blood spot samples was 6 pieces(Φ=2 mm).The average heterozygous peak height ratio across all locus was 0.86.The standard deviation of allelic ladder for 10 runs was within 0.3 bp.The size differences between allele and corresponding allele in allelic ladder was within 0.5 bp.Conclusion Microfluidic chip amplification system enable providing reliable InDel profiles from buccal swabs,dried blood spot,dried saliva spot samples and cigarette butt with accurately ancestral inference.It is possible to yield“sample-in result-out”results of InDel typing in 2 h.This system can meet the requirements of on-site testing and can be used for forensic casework.