金黄色葡萄球菌肠毒素D产毒基因的序列分析

龙军; 陈清; 俞守义

金黄色葡萄球菌肠毒素D产毒基因的序列分析

Amplification and cloning of staphylococcus aureus enterotox in D(SED) gene

摘要

摘要:
目的构建金黄色葡萄球菌肠素D(SED)毒力蛋白基因重组表达质粒。
方法根据SED已知序列, 设计合成一对引物, 用PCR方法从金黄色葡萄球菌毒力质粒上扩增出基因片段, 克隆到原核表达质粒PET32中, 转化大肠埃希菌JM109感受态细胞, 经酶切和PCR鉴定, 然后进行测序。
结果 PCR扩增产物大小为317bp; 重组质粒经双酶切和PCR鉴定表明已正确重组; 测序结果与己知序列基本吻合。
结论国内首次克隆了金黄色葡萄球菌肠毒素D产毒基因, 为下一步研究发病机制奠定基础。

Abstract:
Objective To construct a recombinant plasmid containing staphylococcus aureus toxic protein gene.
Methods A couple of primers were designed for PCR according to the known sequence of SED.The gene obtained by amplification from plasmid DNA of staphylococcus aureus by PCR technique was cloned into plamid of p ET32a directionally.The recombinant plasmid p ET32a was transferred into competent J M109.The recombinants were screened and identified by restriction analysis and PCR, the cloned gene was sequened.
Results The size of amplified PCR products was 317bp.The correct recombinant plasmid p ET32a was isolated and confirmed by restriction analysis and PCR, DNA sequencing showed the DNA sequence of the cloned gene was the same as the published sequence.
Conclusions The SED toxic gene was first successfully amplified and cloned into plasmid PET32a in our country.It provided the basic material for studying the pathogenesis.

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