Objective: To construct a three plasmids lentiviral vector containing canine coagulation factor IX (cFIX) gene with ubiquinone promoter (PUB) and observe the expression of cFIX gene.
Methods: Lentivirus was generated by transient three-plasmid transfection, namely, the VSV-G envelope expression cassette, the delta NRF packaging plasmid and the PTK 164 plasmid. Viral particles were used to infect the target cell, third passage mesenchymal stem cells (MSCs) and 293T cell respectively at MOI 3: 1. The cFIX activity was detected in cultured cells with one-stage clotting assay.
Results: The MSCs were obtained in vitro. The lentivirus infected MSCs and 293T cells all expressed the active factor IX with the activity of (26.30 +/- 2.10)% and (19.70 +/- 1.53)%, respectively, which are significantly higher than that of control (1.00 +/- 0.05)%.
Conclusions: The lentiviral vector of three plasmids with ubiquinone promoter (PUB) was constructed and can transfect the MSCs and 293T cells.