细菌总RNA提取方法的比较

Comparison of Different Methods for Total Bacteria RNA extraction

采用改良前后的Trizol法,RNAiso plus法、Qiagen试剂盒法以及Triton X-100法,从大肠杆菌(Escherichiacoli),枯草芽孢杆菌(Bacillus subtilis),乳酸乳球菌(Lactococcuslactis)3株细菌中提取RNA。用琼脂糖凝胶电泳和核酸浓度测定仪检测总RNA的提取质量,并用RT-PCR(ReverseTranscription Polymerase Chain Reaction)对RNA的质量进行验证。结果表明:RNAiso plus法对3株菌的总RNA提取均有很好的效果。Trizol法和Qiagen试剂盒法对于E.coli总RNA的提取效果不错,但对其他2株菌的提取效果均不明显,并且都伴有DNA污染。上述几种方法提取出的RNA样品中均含有23S和16S rRNA。L.lactis更适合用Triton X-100法提取总RNA。改良后的Trizol法和RNAiso plus法与Triton X-100法的原理基本相同,都是采用加热的方法,更彻底地裂解细菌细胞,而且得到的RNA中不含有rRNA和tRNA,只保留了mRNA,能更好地用于后续的实验研究。

In this research, RNA was isolated from three bacterial strains, （Escherichia coli, Bacillus subtilis and Lactococcuslactis） by using modified and un-modified Trizol and RNAiso plus methods, Triton X-100 method and Qiagen method. The quality and integrity of the extracted RNA samples were determined with agarose gel electrophoresis, nucleic acid detector and RT-PCR detection. The results demonstrate that un-modified RNAiso plus method can effectively collect high-quality RNA from the three bacterial strains. Qiagen method and tin-modified Trizol method showed good efficiency when isolating total RNAfrom E.coli but other strains. In addition, the RNA samples also contain DNA contamination. L. lactis was more suitable for using Triton X-100 method. With the same principle, modified Trizol method, modified RNAiso plus method and Triton X-100 method reported here can be employed for extraction of RNA that are free from 16S and 23S rRNA and provide simple, rapid and effective tools for the isolation of high-quality RNA appropriate for downstream molecular experiments