BTG2基因重组慢病毒载体的构建及其在肺癌A549细胞中的表达与功能

目的 构建重组慢病毒载体pCDH-BTG2,并检测其转染肺癌A549细胞后的表达与功能. 方法 通过PCR技术以含B细胞转位基因(BTG2)的cDNA克隆质粒(HsCD00330081)为模板获得BTG2基因片段,将此片段克隆至慢病毒载体pCDH-CMV-MCS-EF1-Puro,形成pCDH-BTG2,转染PC3细胞检测BTG2的表达;将重组慢病毒载体质粒pCDH-BTG2包装成重组慢病毒颗粒,感染肺癌A549细胞后,通过蛋白免疫印迹法检测BTG2的表达,细胞计数法检测BTG2对A549细胞生长的影响. 结果 pCDH-BTG2重组质粒构建成功,转染PC3细胞后可检测BTG2蛋白的表达,pCDH-BTG2包装成慢病毒感染A549细胞后,显著抑制A549细胞的生长. 结论 成功构建了重组慢病毒载体pCDH-BTG2,并发现BTG2抑制A549细胞的生长,为研究BTG2的功能奠定了基础,同时为肺癌的药物研究提供了靶点.

Construction of Recombinant Lentiviral Vector pCDH-BTG2 and Its Expression and Function in A549 Cells

ObjectiveTo construct recombinant lentiviral vector pCDH-BTG2 and detect its expression and function in A549 cells.MethodscDNA clone plasmid (HsCD00330081) was used as template to amplify the fragment BTG2 by PCR.The fragment was cloned into pCDH-CMV-MCS-EF1-Puro to generate recombinant lentiviral vector pCDH-BTG2.PC3 cells were transfected with pCDH-BTG2 and its expression was detected by Western-blot.Recombinant lentivirus was produced by 293T cells following co-transfection of pCDH-BTG2 with the packaging plasmids.The resulting recombinant lentivirus Lenti-BTG2 was then used to infect A549 cells.BTG2 expression was detected by Western-blot analysis, and cell growth assays were performed to check the effect of BTG2 on A549 cells.ResultsThe recombinant lentiviral vector pCDH-BTG2 was successfully constructed and could be detected by Western-blot after transfecting PC3 cell.Lentivirus Lenti-BTG2 was successfully constructed and could infect A549 cells.Expression of BTG2 obviously inhibited cell growth of A549 cells.ConclusionThe lentiviral vector pCDH-BTG2 was successfully constructed, which lays the foundation for the research of BTG2 function.We also found that BTG2 inhibits lung cancer cell growth, which provides a potential therapeutic agent for lung cancer.