动物造模,慢病毒载体 z-bo 2020-09-09 381

　　Objective: To observe the effect of lentiviral vector-mediated HCV receptor gene OCLN/CD81 transfection on the biological characteristics and gene expression of sh bone marrow mesenchymal stem cells in vitro

　　Method: Isolate and identify the preserved Tupai BM-MSC in this room, construct a lentiviral vector containing human OCLN/CD81 gene, and transfect the CD81 and OCLN genes into BM-MSC respectively. Use an inverted fluorescence microscope and flow cytometer to detect the transfection efficiency. Whether to use CCK8 method to detect the cell proliferation of BM-MSC after transfection? Whether to induce osteoblastic adipocyte differentiation to detect its multidirectional potential? After transfection, real-time fluorescence and WB methods are used to detect HCV receptor (OC/CD81) gene mRNA and protein expression BM-MSCs stem gene expression?

　　Result: After transfection with a lentiviral vector containing human OCLN/CD81 gene with MOI of 100, the transfection efficiency of LV-CD81 reached 99.4%, and the transfection efficiency of LVOCLN was 22.6%? The cell proliferation trend is similar to that of untransfected BM-MSC, but the transfected BM-MSC can differentiate into osteoblasts, but the ability is weaker? The expression of stem gene NANOG mRNA increased, the difference was statistically significant (P\u003c0.05), and the expression of LIN28A mRNA decreased, the difference was significant (P\u003c0.05)? Can pepper BM-MSC express effectively transferred foreign genes after transfection?

　　Conclusion: The constructed lentiviral vector based on the human OCLN/CD81 gene can successfully transfect Tupai BM-MSC and effectively express the introduced gene. Does transfection have a special effect on the multi-directional differentiation potential of BM-MSC?