冷诱导RNA结合蛋白(CIRP)过表达载体的构建及鉴定  

Construction and identification of over-expressing vector of cold inducible RNA-binding protein(CIRP)

作　　者：李静辉[1] 孟宇[1] 李玉恒[1] 李昌盛[1] 杨焕民[1] 李士泽[1,2] 

机构地区：[1]黑龙江八一农垦大学动物科技学院,大庆163319 [2]黑龙江八一农垦大学食品学院,大庆163319

出　　处：《应用与环境生物学报》2015年第2期381-384,共4页Chinese Journal of Applied and Environmental Biology

基　　金：国家自然科学基金项目(31272524);农业部948计划重点项目(2011-G35);黑龙江省自然科学基金项目(C201103);黑龙江八一农垦大学研究生创新实验项目(YJSCX2014-Y25)资助~~

摘　　要：为构建携带冷诱导RNA结合蛋白(Cold inducible RNA-binding protein,CIRP)的过表达载体,首先从4℃条件下冷处理8 h的SD大鼠睾丸组织中获得CIRP基因的c DNA序列,然后扩增CIRP基因的编码区,将其克隆到真核表达载体p Lenti6/V5中,酶切鉴定并测序.将克隆成功的质粒转染HEK293T细胞,用Western blot检测CIRP的表达水平.成功扩增CIRP编码区,并将其克隆至载体p Lenti6/V5中;当将CIRP过表达载体转染HEK293T细胞后,Western blot检测结果显示CIRP蛋白表达水平明显增加(P<0.01).本研究成功构建了CIRP过表达载体,为进一步研究CIRP的作用机制提供了基础工具.To provide a basis for further research of cold inducible RNA-binding protein （CIRP） function, this studyconstructed an over-expression vector of CIRP. After the CIRP cDNA sequencing of testicular tissue of SD rats was obtained with cold treatment at the temperature of 4 ℃, the coding region of CIRP gene was amplified and then cloned into the eukaryotic expression vector pLenti6/V5, followed by endonuclease digestion and sequencing. The successfully cloned plasmid was transfected into HEK293T cells, and CIRP expression was determined by Western blot. The coding region of CIRP gene was successfully amplified, and cloned into the pLenti6/V5. After transfected with CIRP over-expression vectors, the expression of CIRP protein was up-regulated in HEK293T cells （P 〈 0.01）. The results indicated that over-expression vector of CIRP can be successfully constructed and determined, providing a basis for further research of CIRP.

关 键 词：冷诱导RNA结合蛋白 过表达 质粒 冷应激 

分 类 号：Q78[生物学—分子生物学]