两个体混合样本比例的PCR-STR检测研究

The examination of DNA sensitivity in mixed sample

目的探讨PCR-STR检测两个体混合样本比例的灵敏度。方法按不同比例将两个体全血、单个核细胞及纯DNA 3种不同的样本进行混合,应用AB I公司生产的profile p lus商品化试剂盒多重PCR扩增后,用AB I3100基因分析仪进行毛细管电泳,检测基因座及其峰面积,并就混合样本中两单个核细胞样本DNA含量百分比与扩增前混合样本比例的关系进行相关分析。结果当样本混合比例为4%+96%时,可明确区分混合样本中两个体的基因型,且扩增前混合细胞百分率与扩增后两者峰面积比值呈直线相关。结论PCR-STR检测两个体混合样本比例的灵敏度有一定范围,并可进行半定量。

Objective To evaluate the sensitivity of the mixed samples using fluorescence labelled multiplex PCR amplification of STR （short tandem repeats）. Methods To mix two samples with three kinds of different specimen modes （ whole human blood, mononuclear cell and refined DNA） according to different proportions, and was amplified by commercial profile plus PCR amplification kit （ABI, American）. The PCR products and fluorescence detection were performed with ABI prism 3100 genetic analyzer. Genemapper 3.1 software was used for size calling and quantification of peak areas. The formula to calculate DNA percentage of two single samples in the mixed sample was based on genotype distinction of two single samples and to investigate whether the DNA percentage and the proportion of pre - amplified samples mixture should show a linear correlation. Results the results of mixed samples of mononuclear cell and DNA showed a concordance. The genotype of the mixed single sample can be distinguished specific when the mixed proportion of two samples was between 4% -96%, and the ratio of the pre - amplified percentage and the post - amplified peak areas of mixed cells exhibited a linear correlation. Conclusion The sensitivity of STR in mixed samples using a fluorescence labelled multiplex PCR method revealed limited restriction, but can be detected by partial quantitation.