水稻生长素结合蛋白ABP1的原核表达及纯化  

Prokaryotic Expression and Purification of the Auxin Binding Protein 1 in Rice

作　　者：龚蓉[1] 黄志刚[1] 王若仲[1] 黄乐宁[1] 萧浪涛[1] 

机构地区：[1]湖南农业大学植物激素与生长发育湖南省重点实验室,湖南长沙410128

出　　处：《激光生物学报》2011年第1期78-82,共5页Acta Laser Biology Sinica

基　　金：国家自然科学基金项目(90817101;30900099);教育部新世纪优秀人才支持计划(NCET-10-0143);湖南省应用基础研究计划项目(2007FJ2003);湖南省教育厅资助科研项目(09K051;09C503)

摘　　要：生长素结合蛋白能够与生长素特异性结合,因而有可能直接被用作生长素免疫分析和生物传感测定中的高特异性、高亲和力识别分子。本研究通过RT-PCR获得水稻生长素结合蛋白1(ABP1)cDNA,将其克隆到原核表达载体pET-32a(+)中,成功构建pET-32a-ABP1重组表达载体。经酶切、PCR及DNA测序鉴定后,将阳性质粒转化表达受体菌E.coil BL21(DE3)。加入异丙基-β-D-硫代半乳糖苷(IPTG)进行诱导后,取样进行SDS-PAGE和Western Blot分析。结果表明成功表达出一个分子量约为40kD的可溶性融合蛋白,并利用Ni^(2+)-NTA亲和柱一步纯化方式得到了ABP1。The auxin binding protein could be used as a highly specific and affinitive recognizing molecule in the immunoassay and biosensors to detect auxin because of its specific binding capability to the auxin. Rice（ Oryza sativa） auxin binding protein I gene obtained by RT-PCR was cloned into the prokaryotie expression vector pET-32a（ ＋ ） to construct the rec.ombinant plasmid pET-32a-ABPI. After digestion, PCR and DNA sequencing identification, the recombinant plasmid was transformed into expressional bacteria E. coil BI21 （ DE3 ） , then the ABPI protein induced by IPTG and purified by affinitive chromatography was expressed successfully and identified by SDS-PAGE and Western Blot. Conclusion： We have suecessed in expressing a soluble fusion protein about 40 kDa in the E. coil BL21 （DE3） and obtained the purified ABP1 protein with Ni2＋ -NTA affinitive column.

关 键 词：水稻 生长素结合蛋白 原核表达 纯化 

分 类 号：Q785[生物学—分子生物学]