PCR检测结核分枝杆菌DNA中不同细菌裂解方法结果观察

An Observation of Results of The Bacterial Splitting Method in Bacillus Tuberclosis DNA by PCR Technique

目的：寻找适合于临床ＰＣＲ检测结核杆菌ＤＮＡ简单、快速、有效、经济的细菌裂解方法。方法：采用１１种细菌裂解剂对相同数量的结核分枝杆菌进行裂解，裂解上清液直接用于ＰＣＲ扩增，结果：１％ＴｒｉｔｏｎＸ－１００，１％ＮＰ４０，１％Ｔｅｗｅｅｎ２０、和１％ＯＰ四种裂解剂的裂解液可直接扩增出结核分枝杆菌ＤＮＡ；ＳＤＳ，ＮａＯＨ，十二烷基肌酸钠等裂解剂的裂解液直接用于ＰＣＲ扩增均为阴性。结论：１％的Ｔｒｉｔｏｎｘ－１００，ＮＰ４０、Ｔｅｗｅｅｎ２０和乳化剂ＯＰ裂解细菌效果好，又不抑制ＰＣＲ反应，适合于作为临床ＰＣＲ检测结核杆菌的裂解剂。

Objective:To explore a simple,rapid,effective and economic bacterial splitting method in Bacillus tuberclosis DNA by PCR technigue.Method:Eleven kinds of bacterial splitting agents were used for degrading the same amount of Bacillus tuberclosis and the supernatants were performed for PCR amplification.Results:1% concentration of TritionX 100, NP 40,Tween20 or OP treated supernatants all can directly amplify the DNA by PCR technique; and the others, such as SDS, NaOH, dodecyl sodium creatinate etc. all gave negative results. Conclusion:1%concentrations of TritonX-100,NP-40,Teween-20 or emulsion OP all give efficient results without inhbiting PCR reaction,which are the splitting agents for Bacillus tuberclosis detection in clinial PCR technigue.