拟南芥酪蛋白激酶1A的原核表达及纯化  

Prokaryotic Expression and Purification of Arabidopsis Casein Kinase 1A

作　　者：郭明[1] 赵小英[1] 刘选明[1] 

机构地区：[1]湖南大学生物学院,中国湖南长沙410082

出　　处：《生命科学研究》2016年第2期131-134,共4页Life Science Research

基　　金：国家自然科学基金资助项目(31171176;30770200);湖南省生物发育工程及新产品研发协同创新中心(20134486)

摘　　要：拟南芥酪蛋白激酶1A(Arabidopsis casein kinase 1A,At CK1A)属于丝氨酸/苏氨酸蛋白激酶家族,其可能参与了蓝光信号转导并调控植物开花时间。通过对At CK1A的全长序列进行生物信息学分析得到了其激酶区;采用RT-PCR的方法从拟南芥c DNA中扩增At CK1A激酶区,并构建至p ET28a原核表达载体;将表达载体导入BL21(DE3)菌株后,经IPTG诱导表达和Ni亲和层析、阳离子交换层析以及凝胶过滤层析纯化,得到高纯度的重组蛋白;最后利用His标签抗体对重组蛋白进行了验证。Arabidopsis casein kinase 1A （AtCK1A）, belongs to the Ser/Thr protein kinase family, is involved in the blue light signaling and flowering time regulation in A rabidopsis. Here, the kinase domain of AtCK1A was isolated according to the bioinformatic analysis of the protein sequence. Then, the AtCK1A kinase domain was amplified by RT-PCR from A rabidopsis thaliana cDNA, and cloned into the prokaryotic expression vector pET28a. The recombinant protein was expressed in BL21 （DE3） through IPTG induction, and purified by Ni affinity chromatography, cation exchange chromatography and gel filtration chromatography. Furthermore, the identity of the recombinant protein was verified by Western-blot using anti-His antibody.

关 键 词：拟南芥 酪蛋白激酶 原核表达 蛋白纯化 

分 类 号：Q946.5[生物学—植物学]