家蝇抗菌肽Domesticin在毕赤酵母中的表达及抑菌活性检测

doi:10.13560/j.cnki.biotech.bull.1985.2018-0611

摘要/Abstract

摘要： 利用毕赤酵母菌株表达家蝇抗菌肽domesticin基因并检测其抑菌活性。克隆家蝇domesticin基因与pPIC9k质粒相连,构建重组表达载体pPIC9K-domesticin,将其电击转化入毕赤酵母KM71中。甲醇诱导后利用Tricine-SDS-PAGE及Western Blot检测融合蛋白的表达,通过最小抑菌浓度测定表达产物的抑菌活性。结果显示家蝇抗菌肽Domesticin在毕赤酵母中成功表达,抑菌实验表明Domesticin对多种细菌具有抑制作用。Domesticin是一种对受试革兰氏阳性菌和革兰氏阴性菌均具有抑菌活性的广谱新型抗菌肽,有望成为新一代抗菌剂。

关键词: 家蝇, 抗菌肽, 毕赤酵母, 真核表达, 抑菌活性

Abstract: This work aims to express the Musca domestica antibacterial peptide Domesticin gene domesticin in Pichia pastoris and determine the antibacterial activity of the expressed product. The cloned domesticin gene fragment was ligated to pPIC9K vector to construct a recombinant expression vector pPIC9K-domesticin,which was transformed into competent Pichia pastoris KM71. Recombinant fusion protein Domesticin was induced with methanol,the expressed product was identified by Tricine-SDS-PAGE and Western Blot,and antibacterial activity was determined by minimal inhibitory concentration. The results indicated that antibacterial peptide Domesticin was successfully expressed in P. pastoris,and Domesticin demonstrated antibacterial activity against many kinds of bacteria. Domesticin is a novel broad spectrum antibacterial peptide that has antibacterial activity against both Gram-positive bacteria and Gram-negative bacteria,which has the capacity to be a new generation antimicrobial agent.

Key words: Musca domestica, antibacterial peptide, Pichia pastoris, eukaryotic expression, antibacterial activity

刘进兰, 杨雪, 李双双, 张玉明, 柳峰松, 唐婷, 李红权. 家蝇抗菌肽Domesticin在毕赤酵母中的表达及抑菌活性检测[J]. 生物技术通报, 2019, 35(2): 109-115.

LIU Jin-lan, YANG Xue, LI Shuang-shuang, ZHANG Yu-ming, LIU Feng-song, TANG Ting, LI Hong-quan. Eukaryotic Expression of Antibacterial Peptide Domesticin from Musca domestica in Pichia Pastoris and Antibacterial Activity Identification[J]. Biotechnology Bulletin, 2019, 35(2): 109-115.

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