comE基因的表达水平与肺炎链球菌的转化率相关性研究

Correlation between comE expression levels and transformation rates of Streptococcus pneumoniae

目的:探讨肺炎链球菌感受态形成关键基因comE的表达对肺炎链球菌转化的影响.方法:构建肺炎链球菌基因comE的缺陷菌株,将野生菌株与缺陷菌株分别进行自然转化和感受态刺激因子(CSP)诱导转化,记录不同时间段的转化率,并采用FQ-PCR技术检测其comE的mRNA表达水平.结果:comE基因缺陷菌株的自然转化和CSP诱导转化率都为0,且检测不到comE的mRNA表达.R6野生菌株的自然转化率在细菌密度A550 nm为0.3左右时最高,comEmRNA的表达水平与其转化率的相关系数为0.244.D39在不同浓度CSP的诱导下都能发生转化,但CSP浓度为100μg/L时的转化率高于10,1000μg/L两个诱导浓度.100μg/L CSP诱导10 min时comEmRNA的表达量较基础水平显著增高(P<0.05).10,1000μg/L CSP诱导时,其不同时间comEmRNA的表达量与其转化率相关性差.100μg/L CSP诱导时,其转化率与0,20 min时comEmRNA表达量的相关系数分别为0.9966和0.9984;与10 min时comEmRNA表达量的相关系数为0.7950.结论:不管是自然转化还是CSP诱导转化,comE的表达都是必需的,但其表达水平与细菌的自然转化率无显著相关性.CSP诱导转化的最佳浓度为100μg/L,此时的转化率与comEmRNA的基础表达量显著相关,而在10,1000μg/L的CSP诱导时不存在这种相关性.

AIM： To investigated the effect of comE on S. pn transformation. METHODS： ComE-defective strains of S. pn were constructed. The wild S. pn strains and their comE-defective derivations were subjected to natural transformation and CSP-induced transformation. Transformation rates were calculated at different time stages and the mRNA expression levels of comE were detected by FQ-PCR. RESULTS： For comE-defective strain, the transformation rate was 0, whether in natural or CSP-induced state, and the mRNA expression level of come was accordingly undetectable. Whereas, for wild tyep S. pn R6, the natural trans- formation rate peaked at the A550 of about 0.3, and the correlation coefficient between come mRNA level and the transformation rate was 0. 244. Similarly, D39 underwent transformation with the induction of different concentrations of CSP. The transformation rate was the highest with CSP concentration of 100μg/L, higher than that of 10 and 1000μg/L. After 10-min induction with CSP at the concentration of 100μg/L, the mRNA level of come was significantly higher than that of basal level （P〈0.05）. When CSP concentration was at 10 and 1000μg/L, comE mRNA levels at different times were not well correlated with corresponding transformation rates. Under the concentration of 100μg/L, the correlation coefficients between comE mRNA expression level and the corresponding transformation rate were 0.9966, 0.7950 and 0.9984at 0, 10 and 20 min, respectively. CONCLUSION： ComE is an essential gene for natural transformation and CSP- induced transformation, but no obvious correlation between the transformation rate and the come mRNA expression level. The optimal concentration of CSP inducing transformation is 100μg/L, at which the transformation rate is notably correlated with the basal comE mRNA level.