山羊流产衣原体ompA和CPAF基因的序列分析及原核表达

Sequence Analysis and Prokaryotic Expression of ompA and CPAF Genes in Chlamydophila abortus of Goats

为了解山羊流产衣原体陕西分离株ompA和CPAF基因的遗传变异情况及制备检测抗原,根据GenBank收录的ompA和CPAF基因序列设计合成特异性引物,分别进行ompA基因和CPAF基因的基因克隆、序列分析及原核表达。结果表明,成功克隆出大小为1053 bp的ompA基因和大小为1056 bp的CPAF基因;核酸序列分析显示ompA和CPAF基因与已报道的山羊流产衣原体ompA和CPAF基因同源性分别为99.54%~99.91%和91%~100%;成功构建了表达ompA和CPAF基因的原核表达菌株,经IPTG诱导后分别表达38.6 ku和38.7 ku的ompA和CPAF重组蛋白,Western blot显示纯化复性的两种重组蛋白均能与山羊流产衣原体阳性血清相结合。试验结果为进一步建立山羊流产衣原体抗体ELISA检测方法奠定了基础。

In order to understand the genetic variation of the ompA and CPAF genes of Chlamydophila abortus and prepare the detection antigens,specific primers were designed and synthesized according to the sequences of the ompA and CPAF genes in GenBank.The gene cloning,sequence analysis and prokaryotic expression of the ompA and CPAF genes were carried out respectively.The results showed that the ompA gene with a size of 1053 bp and the CPAF gene with a size of 1056 bp were successfully cloned.Nucleic acid sequence analysis showed that the homologies of ompA and CPAF genes were 99.54%-99.91%and 91%-100%,respectively,with the reported ompA and CPAF genes of C.abortus.Prokaryotic expression strains of ompA and CPAF genes were successfully constructed.After IPTG induction,38.6 ku and 38.7 ku OmpA and CPAF recombinant proteins were expressed,respectively.Western blot showed that the purified and refolded recombinant protein can be combined with goat C.abortus positive serum.The results of this experiment laid the foundation for the further establishment of rapid antibody ELISA detection methods.