人ERK2基因对人视网膜母细胞瘤WERI-Rb-1细胞生长的影响

摘要

Objective:To construct the eukaryotic expression vector of human extracellular signal regulated kinase 2(ERK2) labeled with Myc and detect its expression and activity in human retinoblastoma cells.Methods:Human ERK2 coding region was amplified from human mammary cDNA library by PCR and cloned into pXJ-40 vector.The recombinant plasmid Myc-ERK2 was transfected into human WERI-Rb-1 cells and the expression was detected by Western blot.The proliferation of WERI-Rb-1 cells was examined by growth curve experiment.Results:Myc-ERK2 eukaryotic expression vector was constructed and verified by double enzyme digestion and nucleic acid sequencing.Human ERK2 protein was expressed in human WERI-Rb-1 cells as was shown by Western blot results and ERK2 protein promoted the proliferation of WERI-Rb-1 cells.Conclusion:The Myc-ERK2 was identified to promote WERI-Rb-1 cell proliferation,which is a fundamental study for ERK2's role in retinoblaastoma.%目的:构建携带Myc标签的人细胞外信号调节激酶2(ERK2)的真核表达载体,表达Myc-ERK2融合蛋白,检测其对人视网膜母细胞瘤细胞生长的影响.方法:采用PCR技术从乳腺文库中扩增人ERK2基因序列,将其插入pXJ-40载体;将重组质粒与空载体转染人视网膜母细胞瘤WERI-Rb-1细胞系后,Western印迹检测表达情况,并进行生长曲线实验.结果:双酶切和测序鉴定表明Myc-ERK2真核表达质粒构建成功;转染WERI-Rb-1细胞后融合蛋白获得表达,生长曲线实验结果表明ERK2可促进肿瘤细胞的生长.结论:携带Myc标签的人ERK2基因的真核表达载体能在人视网膜母细胞瘤WERI-Rb-1细胞中表达,且能促进该细胞的生长.本实验为进一步研究ERK2在肿瘤尤其是眼恶性肿瘤中的功能奠定了基础.