抗菌肽Omiganan的克隆、表达和抑菌活性  

Cloning,Expressing and Antibacterial Activity of Omiganan Antimicrobial Peptide

作　　者：谢昆[1,2] 李密杰 蒋成砚 鲁海菊[1] 孔琼[1] 何超[1] 沈登荣[1] XIE Kun;LI Mijie;JIANG Chengyan;LU Haiju;KONG Qiong;HE Chao;SHEN Dengrong(College of Life Science and Technology of Honghe University,Mengzi 661199, China;Key Laboratory for Research and Utilization of Characteristic Biological Resources in Southern Yunnan, Mengzi 661199, China)

机构地区：[1]红河学院生命科学与技术学院,云南蒙自661199 [2]云南省高校滇南特色生物资源研究与利用重点实验室,云南蒙自661199

出　　处：《华北农学报》2021年第2期62-67,共6页Acta Agriculturae Boreali-Sinica

基　　金：国家自然科学基金项目(31760638);红河学院生物学重点建设学科资助(SZ1520)。

摘　　要：为获得Omiganan抗菌肽并研究其抑菌活性,根据Omiganan抗菌肽的一级结构(NH2-ILRWPWWPWRRK-COOH),通过基因工程技术,参考大肠杆菌和毕赤酵母偏爱密码子原则,分别获得Omiganan核苷酸序列,再以Omiganan核苷酸序列为模板,设计特异性引物,通过PCR技术扩增Omiganan基因,分别与pET32a表达载体和pPIC9K连接构建pET32a-Omiganan原核表达载体和pPIC9K-Omiganan毕赤酵母表达载体,对含pET32a-Omiganan重组质粒的菌株分别采用IPTG 25℃,IPTG 37℃2种温度和自诱导方式表达获得Omiganan重组蛋白,对含pPIC9K-Omiganan毕赤酵母菌株采用1%甲醇诱导获得Omiganan抗菌肽。结果表明,自诱导14 h后Omiganan重组蛋白的表达量最高,IPTG 25℃,IPTG 37℃2种温度皆能诱导Omiganan重组蛋白的表达,但两者之间无显著差异。通过毕赤酵母表达的Omiganan抗菌肽对大肠杆菌和金黄色葡萄球菌均具有很强的抑菌活性。In order to obtain antimicrobial peptide Omiganan and research its antibacterial activity.In this research,the nucleotide sequence of Omiganan was obtained respectively by genetic engineering technology according to E.coli and Pichia pastoris codons preference and the primary structure of Omiganan antimicrobial peptide(NH2-ILRWPWWPWRRK-COOH),following specific primers were designed to amplify Omiganan gene by PCR.And then the pET32a-Omiganan prokaryotic expression vector and pPIC9K-Omiganan Pichia pastoris expression vector were constructed,the Omiganan recombinant protein was expressed by IPTG 25℃and IPTG 37℃different temperature and self-induction expressing system in E.coli.The Omiganan antimicrobial peptide was expressed by 1%concentration methanol in Pichia pastoris.The results showed that the expression of Omiganan recombinant protein was the highest after induction for 14 h,while there was no significant difference between different IPTG temperature.Antibacterial experiment showed the Omiganan antimicrobial peptide Omiganan expressed by Pichia expression system had strong activity against E.coli and Staphylococcus aureus.

关 键 词：Omiganan 抗菌肽 克隆 表达 抑菌活性 

分 类 号：Q78[生物学—分子生物学]