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【目的】耻垢分枝杆菌(Mycobacterium smegmatis mc2155,mc2155)MSMEG_6281为结核分枝杆菌自溶素Rv3717的同源蛋白,通过建立过表达MSMEG_6281的耻垢分枝杆菌菌株,推测该蛋白对耻垢分枝杆菌肽聚糖代谢的影响。【方法】利用RT-PCR方法检测乙胺丁醇(Ethambutol,EMB)作用后MSMEG_6281基因的表达变化;以耻垢分枝杆菌基因组DNA为模板,采用PCR技术克隆MSMEG_6281基因,构建分枝杆菌表达质粒p VV16-MSMEG_6281,进一步建立MSMEG_6281过表达的耻垢分枝杆菌菌株;利用生长曲线检测MSMEG_6281过表达对耻垢分枝杆菌生长的影响;利用扫描电子显微镜分析MSMEG_6281过表达引起的耻垢分枝杆菌形态变化。【结果】EMB处理引起MSMEG_6281基因表达上调;构建了过表达MSMGE_6281的耻垢分枝杆菌菌株(mc2155/p VV16-MSMEG_6281);过表达MSMGE_6281的耻垢分枝杆菌生长缓慢,菌体形态由短杆状转变为长杆状。【结论】MSMGE_6281的过表达可改变耻垢分枝杆菌形态。MSMGE_6281的功能与细胞壁肽聚糖水解相关,在mc2155细胞壁形态维持方面发挥重要作用。
【Objective】 Mycobacterium smegmatis mc2155 (mc2155) MSMEG_6281 is a homologous protein of Mycobacterium tuberculosis autolysin Rv3717. By establishing a M. smegmatis over-expression strain MSMEG_6281, Effect of mycobacterial peptidoglycan metabolism. 【Methods】 RT-PCR was used to detect the expression of MSMEG_6281 after treatment with ethambutol (EMB). The mycobacterium smegmatis genomic DNA was used as a template to clone the gene of MSMEG_6281 by PCR and to construct the mycobacterial expression plasmid pVV16-MSMEG_6281 to further establish the M. smegmatis over-expression strain MSMEG_6281. The growth curve was used to detect the effect of MSMEG_6281 overexpression on the growth of M. smegmatis. The expression of M. smegmatis caused by MSMEG_6281 overexpression was analyzed by scanning electron microscopy Morphological changes. 【Results】 The results showed that EMB treatment resulted in the up-regulation of MSMEG_6281 gene expression. Mycobacterium smegmatis over-expression MSMGE_6281 (mc2155 / p VV16-MSMEG_6281) was constructed. Mycobacterium smegmatis over-expressed MSMGE_6281 grew slowly, Shape into a long rod. 【Conclusion】 Overexpression of MSMGE_6281 can change the morphology of Mycobacterium smegmatis. The function of MSMGE_6281 is related to cell wall peptidoglycan hydrolysis and plays an important role in the maintenance of mc2155 cell wall morphology.