Cloning of human interleukin 10 and construction of retrovirus vectors

Objective To clone and sequence human interleukin-10 gene and construct retrovirus vectors.Methods Concanavalin A-activated mononuclear cells from peripheral blood were cultured for total RNA extraction.With the use of random primers,cDNA was synthesized and then PCR was per- fored using the specific human interleukin-10 primers to obtain human IL-10 gene fragment.Human IL- 10 was inserted into MSCV vectors with double digestion.Then the recombinant vectors were transfected into PT67 package cells by lipofectin method and screened by G418 selection medium.Results A 534 bp special gene fragment was amplified with RT-PCR and sequence analysis revealed 100% homology to published data in GeneBank.Human IL-10 gene has been inserted into MSCV vectors and virus solution secreting hIL-10 obtained.Conclusion Human interleukin-10 cDNA open reading frame (ORF) was successfully cloned and MSCV-hIL-10 was constructed.The condensed virus solution with high titration of secretion hIL-10 was acquired.