金黄色葡萄球菌tst-1基因敲除菌株的构建

Construction of tst-1-deletion mutant of Staphylococcus aureus

抽提金黄色葡萄球菌834菌株的基因组DNA,PCR克隆扩增tst-1及tst-1的上、下游基因,通过将tst-1上、下游基因分别重组到载体质粒p AULA中,形成同源重组质粒p AULA-Δtst-1,将p AULA-Δtst-1电转入细菌内,进行同源重组,以PCR、Western blot鉴定tst-1基因敲除菌株无tst-1基因片段,且无TSST-1蛋白表达,表明已成功构建金黄色葡萄球菌tst-1基因的敲除菌株。

Abstract The genomic DNA of Staphy＇locoecus aureus strain 834 was extracted. The tst-I gone and it ＇ s upstzveam, downstream genes of S. aureus were amplified with PCR and cloned into plasmid pCRI1. The upstream, downstream genes of tst-1 gene were inserted into plasmid pAULA to form homogenous recombinant plasmid pAULA-Atst-1. The pAULA-Atst-1 was electroporated into S. aureus to carry out homogenous recombination. The tst-l-deletion mutant of S. aureus was characterized with PCR, Western blot for non-tst-1 fragment, no TSST-1 protein expression was found, suggested that the constnlction of tst-l-deletion mutant of S. aureus was successful.