毛白杨木葡聚糖内转糖苷酶/水解酶PtoXTH35原核可溶性表达方法研究

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英文篇名：Study on prokaryotic soluble expression method of xyloglucan endotransglycosylase/hydrolase Pto XTH35 in Populus tomentosa 作者：李钰华 ; 李爽 ; 盖颖 英文作者：Li Yuhua; 关键词：毛白杨 ; 木葡聚糖内转糖苷酶/水解酶 ; 原核表达 ; 包涵体 ; 可溶性蛋白 中文刊名：江苏农业科学 英文刊名：Jiangsu Agricultural Sciences 机构：北京林业大学生物科学与技术学院/林木育种国家工程实验室; 出版日期：2019-05-07 18:05 出版单位：江苏农业科学 年：2019 期：08 基金：国家自然科学基金(编号:31300498) 语种：中文; 页：41-44 页数：4 CN：32-1214/S ISSN：1002-1302 分类号：S792.117

摘要

从毛白杨中克隆得到木葡聚糖内转糖苷酶/水解酶PtoXTH35基因,根据SignalIP在线软件预测蛋白信号肽,并分别构建至pET28a、pET32a、pGEX-4t-1、pMAL-c5e、pET 43.1a等5种原核表达载体,经双酶切和测序验证后转化至BL21(DE3)感受态,使用终浓度为0.4 mmol/L的异丙基-β-D-硫代吡喃半乳糖苷(IPTG)诱导其表达目的蛋白。经SDS-PAGE检测,结果表明5种载体均可表达目的蛋白,pET28a、pET32a、pGEX-4t-1等3种载体所表达蛋白均以包涵体的形式存在,而pMAL-c5e和pET 43.1a载体携带蛋白表达量高并且所表达蛋白50%为可溶性蛋白,实现了毛白杨PtoXTH35在原核表达系统中的高效可溶性表达,该试验为后续进一步研究毛白杨PtoXTH35的体外功能奠定了基础。
        

引文

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