法医学杂志

法医学杂志 ›› 2012, Vol. 28 ›› Issue (2): 123-125.DOI: 10.3969/j.issn.1004-5619.2012.02.012

• 技术与应用 • 上一篇    下一篇

低体积PCR扩增用于单细胞分离和检验

韩俊萍1，李彩霞2，3，严  红2，朱  典4，李艮平4，胡  兰2，3   

  1. （1. 中国人民公安大学，北京 100038； 2. 公安部物证鉴定中心，北京 100038； 3. 法医遗传学公安部重点实验室，北京 100038； 4. 重庆医科大学，重庆 400016）
  • 发布日期:2012-04-25 出版日期:2012-04-28
  • 通讯作者: 通信作者：胡兰，女，博士，主任法医师，主要从事法医遗传学研究；E-mail：hulan328@yahoo.com
  • 作者简介:韩俊萍（1986—），女，山西太原人，硕士研究生，主要从事法医遗传学研究；E-mail：pgww19861025@163.com

Low Volume Amplification in Single Cell Separation and Inspection

HAN JUN-PING1, LI CAI-XIA2,3, YAN HONG2, ZHU DIAN4, LI GEN-PING4, HU LAN2,3   

  1. （1. Chinese People’s Public Security University, Beijing 100038, China; 2. Institute of Forensic Science, Ministry of Public Security, P.R.China, Beijing 100038, China; 3. Key laboratory of Forensic Genetics, Ministry of Public Security, Beijing 100038, China; 4. Chongqing Medical University, Chongqing 400016, China）
  • Online:2012-04-25 Published:2012-04-28

被引次数

13

摘要： 目的 优化在单细胞分离检验中应用0.2 mL试管做低体积扩增载体的最佳反应条件。 方法 制备理想口腔上皮细胞悬液，用0.2 mL试管分别收集捕获到的5、10个细胞，设置蛋白酶K添加、PCR反应金牌酶用量、循环次数3组条件，用Identifiler?誖Plus试剂盒进行复合扩增，比较各组的检出率、等位基因丢失率和非特异性扩增情况。 结果 用0.2 mL试管做低体积扩增中，加蛋白酶K裂解、PCR反应金牌酶0.4 μL、PCR反应32个循环，这3个条件的检出率较高，等位基因丢失率较低。 结论 在单细胞分离检验中采用0.2 mL试管进行低体积扩增，可以作为芯片-低体积扩增的有效补充手段。

关键词: 法医遗传学, 核酸扩增技术, 单细胞

Abstract: Objective To establish optimal amplification conditions with the application of 0.2 mL test tube in single cell separation and inspection. Methods Oral epithelium cell suspension was prepared. Five or ten cells were collected with 0.2 mL test tube. Then DNA were amplified with Identifiler?誖Plus kit in three different conditions in which the proteinase K addition, gold enzyme concentration in PCR reaction, and PCR reaction cycles were adjusted separately. Finally the detection rate, allelic dropout rate and artificial alleles were compared among the groups. Results In these 3 different conditions： addition of proteinase K, addition of 0.4 μL gold enzyme in PCR reaction, and use of 32 cycles, the detection rate was higher and allelic dropout rate was lower than the other conditions. Conclusion In single cell separation and inspection, the usage of 0.2 mL test tube could be an effective supplement to chip-low volume amplification.

Key words: forensic genetics, nucleic acid amplification techniques, single cell

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