磁珠法自动化纯化现场检材DNA方法研究

The study for the DNA case work sample purification by the automated DNA extraction system with magnetic beads

目的利用TE-MAGS在TECAN工作站上结合磁珠试剂盒,建立自动化工作站批量纯化现场检材DNA的方法,并探讨其在法医物证检案中的应用。方法灵敏度测试:标准品使用0.1ng/μL 9947A,用200μL TES稀释制备DNA总量0.1ng^1ng共10种的标准样品,采用本文方法提取纯化,使用IdentifilerTM试剂盒扩增,用3130XL型测序仪检测,Gene Mapper ID-X分析,分析STR图谱质量;纯化能力测试:在1ng总量的标准样品中加入腐殖酸、血红素,采用本文方法提取纯化、扩增检测,分析STR图谱质量;实际案件应用对比:收集304份现场检材,分别采用本方法和硅珠法进行提取纯化,经扩增检测,统计对比两种提取纯化方法 STR分型成功率。结果灵敏度测试:0.1ng^0.2ng总量标准样品提取的DNA模板,扩增后可检测到部分基因座STR图谱,0.3ng^1ng总量标准样品提取的DNA模板,扩增后可以得到完整的STR图谱;纯化能力测试:对混合有一定浓度的腐殖酸、血红素的标准样品的提取产物检测图谱未见明显抑制;实际案件应用对比测试:304份现场检材工作站磁珠法检出成功率(50%)高于硅珠法(40.8%)。结论本文所建立的方法缓冲范围较大,回收率高,纯化能力强,提取产物STR分型成功率高,适合现场检材批量化DNA检验。

Objective To evaluate the forensic application of TE-MAGS technology based on magnetic beads kit on TECAN pipetting platform and establish the automated DNA extraction system of case work samples. Methods Sensitivity test： 10 different DNA samples from 0.1ng to 1ng were prepared with a commercial standard DNA 9947A diluted into 200μL TES. DNA samples were purified by the TE-MAGS technology automatically on the TECAN pipetting platform and then typed using the IdentifilerTM Kit and get the profile of STR with the software GeneMapper ID-X; the power of purification was tested with a trail that purified 1ng DNA mixed with humus acid and hemachrome. Comparative test： 304 casework samples were divided into two purified by TE-MAGS technology and silicon beads respectively to compare the power of purification and the possibility of forensic utility. Results Sensitivity test： 0.3ng and more imported DNA can obtain a good quality of DNA profile compared to the lower imported DNA with dropout of STR peaks （0.1ng and 0.2ng）. The power of purification of the TE-MAGS technology was not affected by humus acid and hemachrome. The comparison result between automatic TE-MAGS technology and manual silicon beads extraction methods from 304 casework samples showed that the former＇s success rate（50%） was higher than the latter（40.8%）. Conclusion The established DNA purification method of TE-MAGS technology automatic DNA extraction system in this study was obviously advantaging at the aspect of success rate, stability, and uniformity and suited to application in the forensic utility future.