Study on cloning and eukaryotic expression of human interleukin 10 gene

Objective To construct a eukaryotic expression vector containing hIL-10 cDNA and express it in CHO cells.Methods hIL-10 cDNA,which was amplified by RT-PCR from human peripheral blood mononuclear cells,was cloned to vector pMD18-T.After the sequence was confirmed,the hIL-10 cDNA was isolated and then was inserted into eukaryotic expression vector pCIneo.The recombinant plasmid pCIneo-hIL-10 was transfected into CHO cells with Lipofectamine Plus~TM Reagent.The expression of hIL-10 in CHO cells was tested by RT-PCR and immunohistochemistry assay; the secretion of hIL-10 in medium was tested by ELASA and western blot assay.Results The sequence of hIL-10 cloned was identical with that was published on GenBank.Mammalian recombinant expression plasmid pCIneo-hIL-10 was constructed.And the transcription and expression of hIL-10 were detected.Conclusion The eukaryotic expression system of hIL-10 was successfully established,which might provide the basis for studying affection of IL-10 in clinics and production of gene engineering medicine.