PCR试验加样过程中无DNA原则控制污染的探讨  

Contamination control in the process of PCR based on “non-DNA ” principle

作　　者：蒋会勇[1] 李海军[1] 韩西群[1] 贺莉[1] 赵彤[1] 

机构地区：[1]南方医科大学南方医院病理科,广州510515

出　　处：《诊断病理学杂志》2005年第1期45-48,i013,共5页Chinese Journal of Diagnostic Pathology

基　　金：广东省科技计划项目 (B3 0 10 1) ;广州市科技计划项目 (2 0 0 2Z3 E40 61)。

摘　　要：目的　探究在PCR操作过程中如何预防污染的一些基本操作方法。方法　首先制定了PCR操作过程中的无DNA原则 ,对实验室的划分 ,操作总原则 ,Eppendorf管开、关 ,有螺旋的离心管开、关 ,装有反应体系Eppendorf管的放置 ,标记反应管的操作 ,移液器的使用方法等进行规范化 ,最大限度地避免DNA污染反应体系。然后通过扩增新鲜扁桃体组织中珠蛋白基因及模拟手套被质粒污染的情况下扩增质粒基因 ,验证该无DNA操作过程对避免污染的有效性。结果　按本实验的规范方法进行操作 ,所有阴性对照均未出现假阳性。结论　采用这种无DNA原则进行PCR操作 ,是控制反应体系污染的一种有效方法。Objective To investigate a way to avoid contamina tion in the process of PCR. Methods A “non-DNA” principle w as formulated in the process of PCR. Through normalization of the processes incl u ding the division of the laboratory, general operating principle, the fine proce ss es of opening and closing of Eppendorf tube and centrifuge tube with a helical c ap, operation of the mark reaction tube and the usage of a sample injector, DNA co ntamination in the reaction system could be avoided to the utmost. Validity of t his “non-DNA” principle was evaluated by amplification of globin gene from fr esh tonsil tissues and plasmid gene on condition that gloves were contaminated b y plasmid. Results Operated on the criterion of the “non-DNA” princip le, we found no false positive in all the negative control groups. Conclusion The “non-DNA” principle is an effective way to avoid contamination in the process of PCR.

关 键 词：多聚酶链反应 标本污染 PCR试验 加样技术 移液器 DNA原则 

分 类 号：R446[医药卫生—诊断学] R394[医药卫生—临床医学]