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- Title
慢病毒介导的HTRA1基因稳定过表达人脑血管平滑肌细胞株的建立.
- Authors
石瑜; 王晓玲; 王敏; 李传芬; 曹秉振
- Abstract
Objective: To construct the lentiviral vector containing HTRA1 as well as its mutant gene(HTRA1-Mut,1091TC) and express them in HBVSMCs. Methods: HTRA1 and HTRA1-Mut gene regional fragment were amplified by RT-PCR and then cloned into the plasmid GV287.Lentiviral plasmid, p Helperl.0, and p Helper2.0 were co-transfected into 293 T cells to obtain recombinant virus. The lentiviral titer was detected. Collected lentivirus was applied to infect human vascular smooth muscle cells. Results: The recombinant lentiviral vectors were constructed and were confirmed by PCR and sequencing analysis. They could efficiently transfect 293 T cells and express in the cells. The lentiviral titer was 2E+8 TU/ml. HBVSMCs produced fluorescent expression after lentivirus transfection, and HTRA1 gene expressed over 80%. Transfected HBVSMCs grew in order without inducing excessive death rate. Conclusion: HTRA1 and HTRA1-Mut recombined lentiviruses with high viral titer were successfully constructed and packaged, and the HTRA1 and HTRA1-Mut gene could be transfected into HBVSMCs with stable and high expression in infected cells, and these cells might be applied for mechanism researches of HTRA1 gene.
- Publication
Progress in Modern Biomedicine, 2016, Vol 16, Issue 12, p2218
- ISSN
1673-6273
- Publication type
Article
- DOI
10.13241/j.cnki.pmb.2016.12.005