产气荚膜梭菌α毒素亚单位基因的表达及表达产物活性分析

Expression and Analysis of Activity of Clostridium Perferingens Alpha Toxin Subunit Gene

含有产气荚膜梭菌α毒素基因序列的克隆载体pMD18TCα ,经BamHⅠ和HindⅢ内切酶双酶切后 ,将α毒素亚单位基因亚克隆到原核表达载体pET30b中 ,构建表达载体pET30bPα。经酶切和测序鉴定 ,α毒素基因中的90 0bp的片段正确插入到表达载体中。重组菌株BL2 1(DE3)pET30bPα经IPTG诱导的表达产物经SDS PAGE和Western_blot分析 ,表达出大小为 38× 10 3的重组蛋白具有和α毒素阳性血清发生特异性的免疫反应。

The subunit gene of %Clostridium perferingens% alpha toxin was subcloned into prokaryotic expression vector pET30b and a recombinant vector pET30bPα was generated.Restriction endonuclease analysis and sequence analysis showed that this sequence about 900 bp was rightly inserted into expression vector.The recombinant vector pET30bPα was transformed into {%E.coli.%} BL21(DE3) strain.SDS-PAGE analysis and Western-blot analysis showed that the expressed products of recombinant strain BL21 (DE3) pET30bPα induced by IPTG was 38×10 3 and had special reaction with alpha toxin antiserum.