魏氏梭菌贵州分离株α-毒素基因的克隆与表达

Cloning and expression of α-toxin gene of Clostridium welchii isolated from Guizhou

提取已鉴定的A型魏氏梭菌Clostridiumwelchii贵州分离株的染色体DNA,经酶切回收分子长2～4kb的DNA片段混合物,将其插入质粒载体pUC19中,并转化至E.coliJM109,通过Amp抗药性和显色反应的选择,再提取阳性重组细菌质粒进行探针检测、酶切分析和PCR鉴定,筛选出含魏氏梭菌α-毒素基因的重组大肠杆菌;经溶血与溶血抑制试验及SDS-聚丙烯酰胺凝胶电泳证实,重组E.coli的表达产物符合α-毒素的生物学特性。

The genomic DNA containing 2-4kb fragments was extracted from the chromosomal DNA of Clostridium welchii in Guizhou province and inserted into pUC19 vecter and expressed in E.coli JM109. The recombinant E.coli was identified by antibiotic and color reactive plates, and the recombined plasmid was detected by α-toxin gene probe, (restriction) endonuclease analysis and PCR. By hemolysis and hemolysis inhibitory test and SDS-PAGE, the product of this gene expressed in E.coli was in conformity with the biological properties of α-toxin.