伤寒沙门菌小RNA T64的功能

Function of small RNA T64 in Salmonella enterica serovar Typhi

目的:探究小RNA T64对伤寒沙门菌(Salmonella enterica serovar Typhi,S.Typhi)表型的影响。方法:用λ-Red重组系统构建小RNA T64的基因缺失变异株,并将携带T64基因序列的重组质粒导入T64缺陷株制备T64回补株,同时将该质粒导入hfq基因缺陷变异株。绘制T64高表达株的生长曲线,观察T64对细菌生长的影响;接种半固体平板观察细菌的动力;结晶紫染色法观察细菌生物膜形成情况。结果:序列分析表明,T64基因缺失变异株构建成功;生长曲线显示,T64高表达促进细菌的生长;与空载体对照株相比,T64高表达株的动力稍减弱;生物膜分析结果显示,T64高表达株生物膜形成能力强于空载体对照株;回补T64基因后,T64缺陷株生物膜形成能力基本恢复,而hfq缺陷株依旧难以形成生物膜。结论:T64能促进细菌生长,且在一定程度上抑制细菌的动力;其有助于S.Typhi形成生物膜,但不足以消除hfq基因缺失对生物膜形成的抑制作用。

Objective: To investigate the effect of the novel small RNA T64 in Salmonella enterica serovar Typhi (S. Typhi) on the phenotype of S. Typhi. Methods:The X-Red recombination system was used to generate the T64 deletion mutant of S. Typhi. A T64 complementary strain was constructed by transferring the recombinant plasmid containing T64 coding region into the deletion mutant. The recombinant plasmid was also transformed to the hfq deletion mutant of S. Typhi. The growth curve of the T64 overexpression strain was drawn to observe the effect of T64 on bacterial growth. The bacterial motility was evaluated on swimming agar plates. Furthermore, the bacterial biofilm formation was analyzed with crystal violet staining.Results:The T64 deletion mutant was constructed successfully, which was confirmed by sequencinganalysis. Bacteria grew faster after J64 overexpression. The motility of the T64 overexpression strain decreaseda little in comparison to the control strain. And the overexpression strain showed an increase in biofilmformation compared with the control strain. After 7^64 gene was complemented, the T64 deletion mutantrestored its ability of biofilm formation, while the hfq deletion mutant still couldnrt form biofilms. Conclusion:The sRNA T64 could promote the growth of S. Typhi and down-regulate the motility in some degree,which may be beneficial to biofilm formation in S. Typhi, but itrs not adequate to erase the repression of biofilm formation by hfq deletion.