Objective      To determine the expression of human clotting factor Ⅸ (hFⅨ) after the recombinant lentiviral vector containing hFⅨ gene infecting human umbilical cord mesenchymal stem cells (hUCMSCs).         Methods      The lentiviral vector GV240-hFⅨ-EGFP containing hFⅨ gene was recombined and transfected with liposome to 293T cells to produce lentiviral particles. hUCMSCs were infected by the plasmid at different multiples of infection (MOI). Fluo

rescence microscopy was used to measure the transfection efficiency

and flow cytometry was employed to confirm the infection. Then hUCMSCs were infected by recombinant lentivirus at the optimal MOI. The expression of hFⅨ at mRNA and protein levels and its activity were studied by RT-PCR

ELISA and A phase method

respectively.       Results      Recombinant lentivirus with a titer of 2×10

8

TU/mL were constructed successfully. When MOI was 10

the infection efficiency of recombinant lentivirus was the highest and reached (92.92±6.32)%. RT-PCR showed hFⅨ gene was only expressed in the hUCMSCs transfected with the lentiviral vector GV240-hFⅨ-EGFP

but not with GV240-EGFP. In 1 d after infection

hUCMSCs effectively expressed hFⅨ

and the expression reached the peak in 5 d after infection. ELISA and A phase method test indicated the peak concentration of hFⅨ reached (87.18±0.86) ng/(10

6

·24 h) and the clotting factor activity of hFⅨ reached (44.3±3.2)% respectively at 5 d. This stable expression maintained at a high level until 29 d after infection.        Conclusion      The hUCMSCs transfected by GV240-hFⅨ-EGFP efficiently express hFⅨ protein at a high level and with biological activity.