C型产气荚膜梭菌β1毒素基因表达

利用PCR技术，从C型产气荚膜梭菌染色体DNA中扩增出β1毒素基因，然后用限制性核酸内切酶BamHI和EcoRI对其进行双酶切处理，回收0．95kb的β1毒素基因片段，最后将其定向克隆在事先经同样内切酶处理的载体pET-28c中相应位点上，转化至受体菌B121(DE3)qh。经BamHI和Eco RI双酶切鉴定和核苷酸序列测定证实，构建的重组质粒pETXBl含有B毒素基因，并且具有正确的基因序列和阅读框架。重组菌株BI21(DE3)(pETXB1)经IPIG诱导后，其表达产物经ELISA检测和SDS-PAGE分析，结果表明重组菌株可以高效表达β1毒素蛋白，该蛋白占菌体总蛋白相对含量的12．24％。

Expression of Beta1 - Toxin Gene from Clostridium perfringens Type C

Beta1-toxin gene was amplified from chromosomal DNA of Clostridium perfringens type C by Polymerase Chain Reaction (PCR),PCR products were cleaved with restriction endonucleases BamHI and EcoRI,then the 0.95kb gene fragments were recovered and inserted into the BamHI/EcoRI site of pET-28c vector.The recombinant plasmid pETXB1 was studied in detail by restriction endonuclease analysis and nucleotide sequencing.The 0.95kb fragment of beta1-toxin gene with positive reading frame was successfully cloned and sequenced.By transformation of BL21(DE3),It got recombinant strain BL21(DE3)(pETXB1).The recombinant strain could produce beta1-toxin protein by ELISA and SDS-PAGE.After recombinant strain was induced by IPTG,its expressed product was about 12.24% of total cellular protein by SDS-PAGE and thin-layer gel scanning analysis.