A型肉毒毒素基因的克隆及序列分析

Cloning and Sequence Analysis of Botulinum Neurotoxin Serotype A

目的:克隆肉毒梭菌(Clostridium botulinum)A型肉毒毒素(BoNTa)编码基因。方法:提取肉毒梭菌国际标准株(62A)基因组DNA,根据肉毒梭菌BoNTa基因(GenBank登录号M30196)序列设计引物,采用LA-PCR方法,扩增出目的基因片段,与pMD18-T载体连接,通过酶切鉴定、测序分析克隆到的A型肉毒毒素基因序列。结果:该基因片段与Genbank中的BoNTa基因序列(GenBank登录号M30196)一致性为100%,预测氨基酸序列一致性为100%。结论:成功克隆肉毒梭菌的A型肉毒毒素基因序列,为肉毒梭菌的快速检测,以及进一步用基因工程方法生产A型肉毒毒素奠定了基础。

Objective-To clone the gene of type A botulinus neurotoxins （BoNTa） from Clostridium botulinum. Methods： The genomic DNA of Clostridium botulinum standard strains （62A） was extracted. The primers were designed and synthesized according to the botulinus neurotoxin gene （GenBank accession number M30196）. The gene of BoNTa was amplified using LA-PCR and ligated to the vector pMD18-T. The recombinant clones were identified by restriction endonucleases and were sequenced. Resultes： The identity of the cloned gene （BoNTa） is 100% with the botulinus neurotoxin gene （Gen- Bank accession number M30196）. Conclusion The cloned gene of type A botulinus neurotoxins from Clostridium botulinum facilitates future study on detection of Clostridium botulinum and expression of the botulinus neurotoxins using gene engineering.