小麦品种“陕253”PDI基因的克隆、原核表达及蛋白纯化  

Isolation,Prokaryotic Expression and Protein Purification of PDI Gene from Wheat Cultivar "Shaan 253"

作　　者：臧闻[1] 董剑[1,2] 高翔[1,2] 陈其皎[1,2] 王延鹏[1] 李志业[1] 史红飞[1] 

机构地区：[1]西北农林科技大学农学院,陕西杨凌712100 [2]陕西省小麦工程技术研究中心/陕西省小麦新品种培育工程研究中心,陕西杨凌712100

出　　处：《麦类作物学报》2011年第5期799-804,共6页Journal of Triticeae Crops

基　　金：国家自然科学基金项目(30900896);陕西省"13115"科技创新工程重大项目(2007ZDKG-01);国家现代农业产业技术体系专项(CARS-3);中央高校基本科研业务费专项资金项目(QN2009007)

摘　　要：为进一步研究控制小麦蛋白二硫键形成的关键蛋白即蛋白质二硫键异构酶(PDI),运用RT-PCR方法克隆出小麦品种"陕253"PDI基因的cDNA序列,基因登陆号为HQ911363。序列分析表明,HQ911363全长为1 539 bp,编码512个氨基酸,含有PDI典型的异构酶活性的催化位点-CGHC-和内质网驻留信号肽-KDEL-。构建该基因的原核表达载体,在宿主菌E.coliBL21(DE3)中经IPTG诱导表达融合蛋白,并对表达蛋白进行了纯化。SDS-PAGE及Western-blot检测证实融合蛋白诱导表达并纯化成功。To further explore the function of PDI gene,a key protein regulating the disulfide formation of proteins in wheat,a cDNA fragment was isolated from wheat cultivar ＂Shaan 253＂ via RT-PCR（GenBank accession No.HQ911363）.The full-length is 1 539 bp,encoding 512 amino acid and containing typical isomerase catalytic site-CGHC-and endoplasmic reticulum retention signal peptide-KDEL-.The expression vector was constructed and transformed into the host bacteria Escherichia coli BL21（DE3）,then the expressed protein induced by IPTG was purified.The fusion protein was successfully expressed and purified by the dual tests of SDS-PAGE and Western-blot.

关 键 词：小麦 蛋白质二硫键异构酶(PDI) 基因克隆 原核表达 蛋白纯化 

分 类 号：S512.1[农业科学—作物学] S330