Construction of the recombinant retroviral vector rRV-vIL-10 and expression in vitro

Objective:To construct retroveral vectors,containing the expression sequence of vIL-10 and to transfect rabbit synoviocyte in vitro with recombinant retrovirus and detect the expression of target genes.Methods:①The primers with restrictive enzyme were designed spot and/amplify target gene by PCR from plasmid including vIL-10 gene was amplified.②The retroviral vector pLXSN of target gene was cloned,and identify the aquired plasmid by sequencing.③Co-transfecting the packaging cell GP-293 with constructed retroviral vector and assistant plasmid pVSVG by calcium phosphate-DNA co-precipitation.The medium containing recombinant virus was collected and titer of virus was determined.④Rabbit synoviocytes was transfected with acquired virus in vitro.Detect the protein expression by cell immunohistochemistry.Results:①The recombinant retrovirus rRV-vIL-10 was successfully constructed.The viral titer reached 5×106 cfu/ml.②vIL-10 gene were transduced into rabbit synoviocytes by recombinant retrovirus in vitro.The protein expression of genes could be detected by cell immunohistochemistry.Conclusion:①The recombinant retrovirus rRV-vIL-10 was successfully constructed.②vIL-10 gene were transduced successfully into the rabbit synoviocytes by retroviral vector in vitro.The transduced synoviocytes can express vIL-10 protein.