cyclinG2表达载体的构建及其对人胃癌细胞生长的作用

Construction of human cyclin G2 gene eukaryotic expression recombinant and it’s effect on the growth of human gastric carcinoma cell

目的:构建包含人cyclinG2基因的真核表达载体,并探讨其对人胃癌细胞体外生长的作用。方法:从POE4细胞总RNA反转录产物中扩增出cyc-linG2cDNA,亚克隆至双顺反子真核表达载体pIRESneo中获得pIRES-G2重组质粒,基因转染SGC-7901细胞,G418筛选阳性克隆,免疫蛋白印迹杂交方法检测cyclinG2蛋白表达情况,平板克隆形成能力和四甲基噻唑蓝(MTT)比色法对转染后细胞的增殖活性进行分析。结果:成功构建包含cyclinG2基因真核重组表达载体pIRES-G2;转染pIRES-G2的SGC-7901细胞克隆的增殖活性明显下降。结论:cyclinG2基因转染后SGC-7901细胞体外增殖能力下降。

OBJECTIVE： To construct human cyclin G2 gene eukaryotic vector and study it＇s effect on human gastric carcinoma cell line growth in vitro. METHODS： The cyclin G2 cDNA fragment was amplified from the reverse transcription products of POE4 cell line by reverse polymerase chain reaction and subcloned into pIRESneo vector to build up recombinant expression plasmid pIRES-G2, identified by colony polymerase chain reaction and sequence analysis. Then it was transferred into SGC-7901cell by cation lipofectin reagent. Cell clones were chosen by G418 and enlarged to culture. The expression level of cyclin G2 protein was examined by using Western blot technique. Cell proliferation ability was tested by clone formation ability and MTT assay. RESULTS： cyclin G2 gene eukaryotic expression vector was constructed successfully, the proliferation ability of cyclin G2 gene transfection anti-clones was lower compared with control group. CONCLUSIONS： cyclin G2 gene could inhibit the growth of SGC-7901. It may be a negative regulator in gastric carcinoma cell cycle regulation.