检验医学与临床2016，Vol.13Issue(18)：2553-2555,3.DOI:10.3969/j.issn.1672-9455.2016.18.001
金黄色葡萄球菌肠毒素B基因在大肠杆菌中的克隆及表达
Cloning and expression of Staphylococcus aureus enterotoxin B gene in Escherichia coli
摘要
Abstract
Objective To amplify the Staphylococcus aureus enterotoxin B (SEB) gene and to construct its prokaryotic expres‐sion vector for conducting induction and expression to lay the foundation for its application research .Methods According to SEB gene sequences in the GenBank ,a pair of specific primer containing BamH Ⅰ and Xho Ⅰ enzyme digestion sites were designed .The SEB genome DNA served as the template for conducting PCR amplification ,then after BamH Ⅰ and Xho Ⅰ dual enzyme digestion , connected with the pET‐28α(+ ) for making corresponding enzyme digestion .Escherichia coli BL21 was transformed .The plasmids were extracted for conducting the dual enzyme digestion identification and sequencing .The fusion protein was expressed by using the IPTG induction .The expressed product was identified by Western blot .Results SEB gene was successfully amplified ,which size was 801bp ,the recombinant PET‐28α(+ )‐SEB dual enzyme digestion identification could find the target fragments ,the sequen‐cing results displayed that SEB was in the correct reading frame ,the sequencing comparative analysis showed that its consistency to nucleotide sequence in the related report was 99% .After IPTG induction ,the fusion protein could be expressed as the inclusion body in the corresponding molecular weight 35 × 103 of pET‐28α(+ )‐SEB/BL21 .The target protein could be detected in the corre‐sponding molecular weight by Western blot .Conclusion SEB gene is cloned and successfully expressed as the inclusion body in Escherichia coli BL21 ,which lays a foundation for staphylococcus aureus SEB application research .关键词
金黄色葡萄球菌/肠毒素B/克隆/表达Key words
Staphylococcus aureus/enterotoxin B/clone/expression引用本文复制引用
李飞,杨镒宇,周珍文,周帅,董慧,黄艳梅,梁秉绍,李娟,龙燕,王洁琳,谢永强..金黄色葡萄球菌肠毒素B基因在大肠杆菌中的克隆及表达[J].检验医学与临床,2016,13(18):2553-2555,3.基金项目
广州市医药科技重点项目（201102A212013）；广东省科技厅项目（2014A020212013）。 （）
