法医学杂志

法医学杂志 ›› 2014, Vol. 30 ›› Issue (5): 342-345.DOI: 10.3969/j.issn.1004-5619.2014.05.006

• 论著 • 上一篇    下一篇

微滴式数字PCR技术用于生物样品种属鉴定和绝对定量

胡  伟，陈荣华，张  晨，安志远，王  斌，平  原   

  1. （上海市公安局物证鉴定中心 法医物证学现场应用技术公安部重点实验室 上海市现场物证重点实验室，上海 200083）
  • 发布日期:2014-10-25 出版日期:2014-10-28
  • 通讯作者: 平原，男，主任法医师，主要从事法医DNA分析技术的应用和研究；E-mail：pingyuan803@126.com
  • 作者简介:胡伟（1974—），男，安徽合肥人，副主任法医师，主要从事法医DNA分析技术的应用和研究；E-mail：hwzhyy1999@ 163.com
  • 基金资助:

    公安部科技强警基础工作专项（2013GABJC004）

Species Identification and Absolute Quantification of Biological Samples by Droplet Digital PCR

HU WEI, CHEN RONG-HUA, ZHANG CHEN, AN ZHI-YUAN, WANG BING, PING YUAN   

  1. （Shanghai Key Laboratory of Crime Scene Evidence, Key Laboratory of Forensic Evidence and Science Technology, Ministry of Public Security, Institute of Forensic Science, Shanghai Public Security Bureau, Shanghai 200083, China）
  • Online:2014-10-25 Published:2014-10-28

被引次数

28

摘要： 目的 运用微滴式数字PCR技术进行生物样品的种属鉴定和绝对定量。 方法 选择人mtDNA两个编码基因ND4和16S rRNA，设计特异性引物与探针，用人来源及常见动物样本验证种属特异性，再用重组质粒和2组共16份人来源生物检材，倍比稀释。使用微滴式数字PCR技术进行种属检验和绝对定量，验证其灵敏度和稳定性。 结果 人重组质粒FAM（ND4）可进行人来源样品的检测，其检测结果与各级稀释梯度基本吻合，微滴式数字PCR技术可以检测出反应体系中低至单拷贝的DNA。 结论 微滴式数字PCR技术可以进行生物样品的种属鉴定和绝对定量，并具有很高的灵敏度和特异性，可应用于日常法医物证检验。

关键词: 法医遗传学, 聚合酶链反应, 种属鉴定, 绝对定量, 微滴式数字PCR

Abstract: Objective To test droplet digital PCR for species identification and absolute quantification of biological sample. Methods Specific primers and probes for human mtDNA encoding gene ND4 and 16S rRNA were designed, and the species-specificity was assessed on DNA samples derived from human and common animals. To determine the sensitivity and stability of droplet digital PCR for species identification and absolute quantification, gradient dilution series of recombinant plasmid and 16 human DNA samples were analyzed. Results Human recombinant plasmid FAM （ND4） could be used in detecting the samples of human. And the results of detecting were consistent with all levels of diluted concentrations. Droplet digital PCR was able to detect low and single copy of target DNA. Conclusion Droplet digital PCR, with high sensitivity and specificity, is fully amenable for species identification and absolute quantification of biological samples, also it can be applied on routine forensic examination.

Key words: forensic genetics, polymerase chain reaction, species identification, absolute quantification, droplet digital PCR

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