适用于结核分枝杆菌RNA抽提的实验方法学研究  

Methodological research on extracting total RNA from tuberculosis bacteria

作　　者：俞垭美 王姗姗[1] 褚丽花 冯宜[1] 葛洪斌 张喆[1] 

机构地区：[1]浙江大学医学院基础医学院感染与免疫研究中心病原生物学系,浙江杭州310058

出　　处：《临床肺科杂志》2016年第4期609-612,共4页Journal of Clinical Pulmonary Medicine

基　　金：浙江省自然科学基金资助(No LY12H19005);浙江大学生科研训练计划项目SRTP第十六期;第十七期资助

摘　　要：目的分枝杆菌的细胞壁结构和成分特殊而复杂,在RNA提取的实验方法学上是否区别于一般的细菌,有待考察。本文使用了异硫氰酸胍提取法与商业化的细菌RNA抽提试剂盒法进行对照,对减活牛型分枝杆菌卡介苗(BCG)进行RNA的抽提,进而确定了适用于分枝杆菌RNA抽提的最优方法。方法分别用异硫氰酸胍提取法和E.Z.N.A.^(TM)Bacterial RNA提取试剂盒从一定量的卡介苗细菌培养物中提取RNA,用Nanodrop2000c检测抽提所得的RNA的浓度和纯度。结果用异硫氰酸胍法从1到30 mg的BCG中均能提取到大量的RNA,而用E.Z.N.A.TMBacterial RNA提取试剂盒几乎未提取到RNA,而且在纯度上前者好于后者。结论异硫氰酸胍的裂解效率远远高于溶菌酶。对于分枝杆菌例如结核分枝杆菌RNA的提取应该使用异硫氰酸胍法。Objective To explore the best method to extract total RNA from tuberculosis bacteria. Methods The E. Z. N. A TM Bacterial RNA Kit and guanidine thiocyanate （GTC） method were used to isolate mycobaeterial RNA from different amount of BCG. The quantity and quality of the isolated RNA were compared. Results The quantity and quality of total RNA extracted by GTC method were both higher than bacterial RNA extraction method. Conclusion The cell lysis efficiency of GTC is higher than that of lysozyme. GTC method is a better method than common bacterial RNA extraction to extract method total RNA from mycobacteria.

关 键 词：结核分枝杆菌 RNA提取 异硫氰酸胍(GTC) 

分 类 号：R440[医药卫生—诊断学]