牛瑟氏泰勒虫P23表面蛋白基因表达条件的优化

Optimization of expression condition of the Cattle Theileria sergenti P23 surface protein gene

将已构建并测序正确的含有牛瑟氏泰勒虫P23表面蛋白基因的pGEX-4T-P23转化菌用IPTG进行诱导表达,经SDS-PAGE电泳可检测到相对分子量为46.0 ku的融合蛋白.根据SDS-PAGE确定融合蛋白的最佳表达条件,结果显示诱导时机和时间是影响表达的主要因素,诱导温度和IPTG浓度次之;确定最佳诱导时机为转接种后2.0 h,最佳诱导温度34℃,最佳诱导时间6.0 h,最适IPTG浓度0.08 mmol/L.表达产物主要以包涵体存在,在优化条件下融合蛋白的表达量经Bandscan5.0软件分析约占菌体总蛋白的31.7%.

Constructed and sequenced pGEX-4T-P23, which contains surface protein gene P23 of Theileria sergenti was induced by IPTG. Detected the fusion protein relative molecular weight was 46 ku by SDS-PAGE. The best expression of condition was identified on the basis of SDS-PAGE detection. The results showed that inducing juncture and inducing time were the main factors affecting the expression. The induced temperature, IPTG concentration is followed. The optimal inducing juncture is 2.0 h after inoculated,the optimal induction time is 6.0 h, the optima/temperature is 34 ℃ ,and the optimal concentration of IPTG is 0.08 mmol/L. Main expression products existed by inclusion bodies. In the optimized expression conditions,the fusion protein account 31.7% of the total bacterium protein by Bandscan5.0 anatysising.