55株芽孢杆菌16S rRNA基因序列测定与系统发育学分析

Identification of 55 Bacillus Strains by Phylogenetic Analysis of 16S rRNA Gene Sequence

采用16S rRNA基因序列分析法对中国工业微生物菌种保藏管理中心(CCIC)保藏的55株枯草芽孢杆菌(Bacillus subtilis)进行复核鉴定。菌株经纯化培养,以改良CTAB法提取总DNA,采用细菌16S rRNA通用引物、TD-PCR方法(touchdown-PCR)进行16S rRNA基因序列扩增,PCR产物纯化后直接进行序列测定,序列经人工校对后用Clustal X进行比对分析,最后用MEGA3.1软件构建系统发育树。系统发育分析结果表明:55株枯草芽孢杆菌中有52株菌种与原鉴定结果一致,有3株菌种与原鉴定结果存在差异,其中2株鉴定结果为巨大芽孢杆菌(B.megaterium),另1株鉴定结果为地衣芽孢杆菌(B.licheniformis)。

We identified 55 Bacillus strains from China Center of Industrial Culture Collection （CICC） by 16S rRNA gene sequence analysis. The genomic DNA was isolated and purified with the improved CTAB methods. The sequences of 16S rRNA gene were amplified by TD-PCR with the bacterium universal primers and the purified PCR products were directly sequenced. The corrected sequences were aligned with Clustal X and the phylegenetic tree was constructed by MEGA3.1. 52 strains support the original identification results, 2 strains belong to Bacillus megaterium, and the other one belongs to Bacillus lecheniformis using the phylogenetic analysis.