包装重组腺病毒Adncam1及其体外转染神经干细胞

Packaging of recombinant adenovirus Adncam1 and Adncam1 transfect neural stem cells in vitro

目的构建腺病毒载体,将目的基因NCAM1转染入神经干细胞(NSCs)。方法制备重组质粒,完成后与大骨架质粒结合对293A细胞转染,包装成重组腺病毒载体Adncam1。对进行两次扩增,并检测滴度值。利用Adncam1转染NSCs,转染后应用QPCR、Western blot对NSCs中的NCAM1进行过表达鉴定。结果Adncam1包装成功,两次扩增后,检测滴度值为1×1010 PFU/m L。Adncam1对NSCs转染效果明显。进行过表达鉴定,QPCR检测Adncam1转染效果为对照转染的15倍以上,Western blot检测转染效果为3倍以上。结论重组腺病毒Ad Ncam1成功的将目的基因NCAM1转入NSCs。

Objective Construct recombinant adenovirus vector, the NCAM1 gene was transfected into the NSCs. Methods The recombinant plasmid was preparated transfected 293A cells become Adncamlwith the backbone plasmid. After two amplification, testing titer value. Using Adncaml transfected with NSCs, and determine whether the gene NCAM1 was over expres- sion in NSCs by QPCR and Western blot. Results Adncaml was packinged successful, after two amplification, The titer value was 1 ~ 101~ PFU/mL. The effect Adncaml transfected with NSCs was obvious. The transfection efficiency was 15 times higher than the control transfected by QPCR, The transfection efficiency was 3 times higher than the control transfected by Western blot. Conclusion Successful the recombinant adenovirus AdNcaml transfected the target gene NCAMI into NSCs.