稳定表达Prohibitin 2蛋白SW620细胞株的建立  

Establishment of a stable SW620 cell strain expressing prohibitin 2 protein

作　　者：武标[1] 徐丽丽[2] 王妍[1] 汤玲[1] 孙文旦[1] 

机构地区：[1]无锡市第二人民医院检验科,江苏省214002 [2]无锡市妇幼保健院

出　　处：《江苏医药》2015年第20期2373-2376,共4页Jiangsu Medical Journal

基　　金：南京医科大学科技发展基金(2013NJMU183)

摘　　要：目的构建prohibitin 2(Phb2)蛋白基因的真核表达载体,并建立稳定表达Phb2蛋白的细胞株。方法扩增Phb2基因全长编码序列,经双酶切后克隆至真核表达载体pcDNA3中,将重组表达质粒pcDNA3-FLAG-Phb2转染至SW620细胞,用G418筛选阳性克隆,Western blot检测Phb2蛋白的表达。结果经测序鉴定,成功构建了Phb2基因的真核表达载体。转染SW620细胞后能够稳定有效的表达Phb2蛋白。结论成功构建了稳定表达Phb2蛋白的细胞株。Objective To construct a eukaryocyte expression vector of prohibitin 2gene,and establish the stable cell strain expressing the protein continuously.Methods PCR method was used to amplify the open reading frame of prohibitin 2from pLenti6/V5-D-TOPO-prohibitin 2-WT plasmid.Then the fragment was inserted into pcDNA3 vector after digested by KpnⅠand EcoRⅠto construct expression vector pcDNA3-prohibitin 2.SW620 cells were transfected with plasmid pcDNA3-prohibitin 2.Then the cells were selected under the pressure of G418 and Western blot was used to detect the expression of prohibitin 2 protein.Results Confirmed by sequencing,a eukaryotic expression vector encoding FLAG-prohibitin 2fusion protein was successfully constructed.FLAGprohibitin 2fusion protein could be expressed stably in SW620 cells after transfected with plasmid pcDNA3-prohibitin 2.Conclusion A stale cell strain over-expressing prohibitin 2 protein is successfully established.

关 键 词：基因表达 PROHIBITIN 2 

分 类 号：R329[医药卫生—人体解剖和组织胚胎学]