生长素结合蛋白AtTIR1和IAA28的原核表达及纯化  

Prokaryotic expression and purification of the auxin-binding protein AtTIR1 and IAA28

作　　者：赵欣[1] 刘会珍[1] 罗为桂[1] 苏益[1] 蔺万煌[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：《湖南农业大学学报（自然科学版）》2013年第6期609-614,共6页Journal of Hunan Agricultural University(Natural Sciences)

基　　金：国家自然科学基金重大研究计划项目(91317312);湖南省高校创新平台开放基金项目(11K032;13K065)

摘　　要：以pGEX–KG为骨架,运用分子克隆方法构建了生长素结合蛋白AtTIR1和IAA28的原核表达载体,通过转化不同表达菌株探索了2种蛋白的诱导表达条件。结果表明:在0.4 mmol/L IPTG诱导下,GST–IAA28融合蛋白在表达菌株Tuner中的表达量最高,其适宜诱导温度为16℃;在0.6 mmol/L IPTG诱导下,BL21(DE3)中GST–IAA28的表达量最高;在各种浓度的IPTG诱导下,Rosetta中的GST–IAA28的产量均不高;在25℃和0.4mmol/L IPTG诱导条件下,Rosetta中的GST–AtTIR1表达总量最高,每克细菌可诱导出约0.2 mg的目标蛋白,但BL21(DE3)菌种中的GST–AtTIR1表达量很低。利用GST SefiroseTM resin亲和树脂对表达的蛋白进行纯化,获得了较纯的AtTIR1和IAA28蛋白。Prokaryotic expression vector of auxin-binding protein AtTIR1 and IAA28 were constructed by gene cloning based on pGEX-KG. The expression induction conditions of two kinds of different expression strains after transfected with the recombined expression vectors were investigated. The results showed that the GST-IAA28 had the highest yields in Tuner under 0.4 mmol/L IPTG and the highest yields in BL21（DE3） at 0.6 mmol/L IPTG~ with 16 ~C the suitable inducing temperature, while the protein yields were low in Rosetta under every concentration of IPTG. GST-AtTIR1 expressed highly in Rosetta at the optimal induced condition of 0.4 mmol/L IPTG and 25 ℃, and about 0.2 mg GST-AtTIR1 was produced in lg bacteria. But GST-AtTIR1 expressed lowly in BL21. Meanwhile, relatively purified GST-AtTIR1 and GST-IAA28 had been obtained through GST SefiroseTM resin.

关 键 词：生长素结合蛋白 AtTIR1基因 IAA28基因 原核表达 蛋白质纯化 

分 类 号：Q786[生物学—分子生物学] Q949.748.3