Objective: The relA gene,a gene associated with the synthesis of rigorous response signaling molecule (p)ppGpp in Burkholderia pseudomallei, was knocked out and its effects on the biological functions of bacterial gro...Objective: The relA gene,a gene associated with the synthesis of rigorous response signaling molecule (p)ppGpp in Burkholderia pseudomallei, was knocked out and its effects on the biological functions of bacterial growth, motility and biofilm formation were investigated. Methods: The plasmids with trimethoprim resistance (TPR) were modified by enzyme digestion and enzyme ligation method. The TPR fragment was linked to the suicide plasmid pK18mobSacB containing SacB sucrose killing gene by Bgl Ⅱ enzyme digestion site, and trimethoprim resistance was obtained Plasmid(TPR-pK18mobSacB);The relA gene of Burkholderia pseudomallei HNBP001 was knocked out by homologous recombinant gene knockout method, and the relA mutant was obtained. The growth, motility and biofilm phenotypes of Burkholderia pseudomallei HNBP001 were compared before and after the deletion of relA gene. Results: The relA mutant was successfully constructed. The growth rate, motility and biofilm formation of ΔrelA decreased. Conclusion: The modified plasmid TPR- pK18mobSacB can improve the knockout efficiency of HNBP001 strain, which can be widely used in the gene knockout of Burkholderia pseudomallei, and provide convenience for the laboratory to study the gene function of Burkholderia pseudomallei at the molecular level;The mutant of the relA gene inhibits the growth, motility and biofilm formation of HNBP001.展开更多
基金Fund Project:Major Science and Technology Program of Hainan Province(No.ZDKJ202003)。
文摘Objective: The relA gene,a gene associated with the synthesis of rigorous response signaling molecule (p)ppGpp in Burkholderia pseudomallei, was knocked out and its effects on the biological functions of bacterial growth, motility and biofilm formation were investigated. Methods: The plasmids with trimethoprim resistance (TPR) were modified by enzyme digestion and enzyme ligation method. The TPR fragment was linked to the suicide plasmid pK18mobSacB containing SacB sucrose killing gene by Bgl Ⅱ enzyme digestion site, and trimethoprim resistance was obtained Plasmid(TPR-pK18mobSacB);The relA gene of Burkholderia pseudomallei HNBP001 was knocked out by homologous recombinant gene knockout method, and the relA mutant was obtained. The growth, motility and biofilm phenotypes of Burkholderia pseudomallei HNBP001 were compared before and after the deletion of relA gene. Results: The relA mutant was successfully constructed. The growth rate, motility and biofilm formation of ΔrelA decreased. Conclusion: The modified plasmid TPR- pK18mobSacB can improve the knockout efficiency of HNBP001 strain, which can be widely used in the gene knockout of Burkholderia pseudomallei, and provide convenience for the laboratory to study the gene function of Burkholderia pseudomallei at the molecular level;The mutant of the relA gene inhibits the growth, motility and biofilm formation of HNBP001.
