医学分子生物学杂志2012，Vol.9Issue(5)：320-325,6.DOI:10.3870/j.issn.1672-8009.2012.h5.002
重组人p66Shc/腺病毒表达载体的构建与表达
Construction and Expression of Human Recombinant p66Shc Adenovirus
摘要
Abstract
Objective To construct the human recombinant p66Shc adenovirus and observe the expression in human umbilical vein endothelial cells (HUVECs) . Methods Human p66Shc gene was amplified by PCR from pcDNA3. lhis-p66Shc and then directly cloned into the linearized adeno-viral vector in vitro. After confirmed by screening and gene sequencing, the linearized recombinant plasmid was transfected into HEK293A cells for packaging. The p66Shc expression in HEK293A cells and HUVECs infected by Adeno X-p66Shc was detected by Western Blot and Immunofluores-cence cytochemistry staining techniques. Results Gene sequencing indicated that pAdeno X-p66Shc contained p66Shc gene. The cytopathic effect was observed in the HEK293A cells at 10 days after pAdeno X -p66Shc transfection. The expression of p66Shc protein was significantly upregulated in HEK293A cells and HUVECs at 48h after Adeno X -p66Shc infection. Conclusion The Adeno-X Adenoviral System3 ( CMV) can be used to efficiently and conveniently construct recombinant adenovirus containing p66Shc gene capable of efficiently infecting HUVECs.关键词
p66Shc/腺病毒/同源重组/脐静脉内皮细胞Key words
p66Shc/ adenoviruses/ homologous recombination/ human umbilical vein endo-thelial cells分类
生物科学引用本文复制引用
徐蓉,胡刚,林雅军,柳江,魏洁,程建华,许晓东,孙洪范..重组人p66Shc/腺病毒表达载体的构建与表达[J].医学分子生物学杂志,2012,9(5):320-325,6.基金项目
国家自然科学基金(No.81001439),天津科技计划应用基础与前沿计划重点项目(No.08JCZDJC16000) （No.81001439）
