MYCT-1基因真核表达载体的构建及鉴定

Construction and identification of an eukariotic expression vector of MYCT-1

目的构建MYCT-1(myc target)基因真核表达载体,观察其转染胃粘膜GES-1细胞系后的表达。方法用RT-PCR法合成MYCT-1 cDNA,克隆至表达载体pcDNA3.1上,测序验证无突变发生。将表达载体转染GES-1细胞,用RT-PCR和Western blot检测MYCT-1基因的表达。结果限制性内切酶酶切和DNA测序证实成功克隆了MYCT-1 cD-NA全长并正确插入了表达载体,在GES-1细胞中检测到MYCT-1 mRNA和蛋白。结论成功构建了MYCT-1真核表达载体,可在GES-1细胞中稳定表达。

Objective To construct an eukariotic expression vector of MYCY-1 and detect its expression after transfected in GES- 1 cells. Methods MYCT- 1 eDNA was synthetized by RT-PCR and then cloned into an expression vector PeDNA3.1. DNA sequencing was performed to avoid any mutation in recombianant vector. The vector peDNA3.1-MYCT-1 was transfected into GES cells by lipnsome and the expression of MYCT-1 was detected by RT-PCR and Western blot. Results The full length cDNA of MYCT-1 cloned into the exoression vector was identified by restricted enzyme digestion and sequencing. MYCT-1 mRNAs and protein were detected in GES-1 ceils. Conclusion An eukariotic efxpression vector of MYCT-I was ssuccessfitlly constructed. The reconbinant vector could expressed in GES-1 cells steadily.