摘要
用PCR从含产气荚膜梭菌α毒素基因的质粒pXETA1中扩增出α毒素基因 ,用NcoI和BamHI双酶切该α毒素基因 ,回收 0 .95kb的α毒素基因片段 ,再用NcoI和BamHI双酶切含产气荚膜梭菌 β毒素基因质粒pXCPAB2 ,与上述回收的α毒素基因片段连接 ,转化至受体菌BL2 1(DE3)中。经NcoI、BamHI、NcoI酶切反应鉴定和核苷酸序列分析证实 ,获得了理想重组质粒pXCPAB2 ,该重组质粒含有α β融合基因。重组菌株BL2 1(DE3) (pXCPAB2 )经IPTG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α β融合蛋白 ,该融合蛋白占菌体总蛋白的 2 2 .14%。
Alpha_toxin gene was amplified from plasmid pXETA1 Containing Closrtidium perfringens alpha_toxin gene by Polymerase Chain Reaction (PCR),PCR products were cleaved with restriction endonucleases NcoI and BamHI and recovered. The 3 terminus of α toxin gene were genetically fused to the 5 terminus of β toxin gene of Closrtidium perfringens. The recombinant plasmid pXCPAB2 was studied in detail by restriction endonuclease analysis and nucleotide sequencing.The results have shown that the recombinant plasmid carried α β fusion gene.By transformation of BL21(DE3),we got recombinant strain BL21(DE3)(pXCPAB2).The recombinant strain could produce α β fusion protein by ELISA and SDS_PAGE.After recombinant stain was induced by IPTG, its expressed product was about 22.14% of total cellular protein by SDS_PAGE and thin_layer gel scanning analysis.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2001年第5期356-359,共4页
Chinese Journal of Preventive Veterinary Medicine
关键词
产气荚膜梭菌
Α毒素
Β毒素
融合基因
基因表达
Closridium perfringens
Alpha_toxin
Beta_toxin
Fusion gene
Gene expression
