抗菌肽Magainin Ⅱ的原核表达与抗菌活性分析  

Prokaryotic expression and antimicrobial activity analysisi of Magainin Ⅱ gene

作　　者：卢艳敏[1,2] 崔海信[1] 王承民[3] 何宏轩[3] 

机构地区：[1]中国农业科学院农业环境与可持续发展研究所,北京100081 [2]衡水学院生命科学系,河北衡水053000 [3]中国科学院动物研究所动物生态与保护生物学院重点实验室,国家野生动物疫病研究中心,北京100101

出　　处：《河北农业大学学报》2011年第5期59-62,68,共5页Journal of Hebei Agricultural University

基　　金：中国科学院绿色农业技术集成与发展中心创新方向项目(Y029891134)

摘　　要：根据Magainin Ⅱ氨基酸序列,选用大肠杆菌偏爱密码子,化学合成了2个DNA片段,利用重叠区互补PCR法得到Magainin Ⅱ的基因序列。构建了融合表达载体pET21b-Magainin Ⅱ,转化大肠杆菌BL21(DE3)pLysS,IPTG诱导表达,表达产物经SDS-PAGE分析,在5.1 kD处可见目的蛋白条带,经Western blot检测表明成功地在大肠杆菌中表达了融合蛋白。诱导温度在20℃,IPTG终浓度为0.4 mmol/L,诱导时间为12 h,融合蛋白表达效果较好,且主要以可溶性形式存在,对纯化得到的融合蛋白进行活性分析,抑菌活性试验表明,表达产物对大肠杆菌和金黄色葡萄球菌均有抑制作用。Two sequences were designed using codon preference of Escherichia coli and synthesized by chemical method according to amino acid sequence of MagaininⅡ.The whole MagaininⅡgene fragment was obtained by overlap extension PCR.The amplified fragment was inserted into pET-21b and the fusion expression vector pET21b-MagaininⅡ was constructed.The recombinant plasmid was transferred into E.coli BL21（DE3） plyS and induced by IPTG.SDS-PAGE and Western-blotting analysis showed that the recombinant had expressed a 5.1 kD specific protein in E.coli.The fusion protein was expressed mainly in a soluble form after induction with 0.4 mmol/L IPTG at 20 ℃for 12 h.The fusion protein was purified and then antibacterial activity was tested.The activity assay demonstrated that the antimicrobial peptide had antibacterial activities against E.coli and Staphylococcus aureus.

关 键 词：MAGAININ Ⅱ 抗菌肽 融合表达 抑菌活性 

分 类 号：S852.6[农业科学—基础兽医学]