Cloning and eukaryotic expression of human interleukin-29 gene

Objective To clone human interleukin-29(IL-29) gene and express in eukaryotic cells.Methods Total RNA was extracted from human peripheral blood mononuclear cells(PBMCs) and reversely transcribed into cDNA as a template with which IL-29 gene was amplified by PCR and inserted into eukaryotic expression vector pPIC9K.The constructed recombinant plasmid pPIC9K-29 was transformed to Pichia pastoris GS115 and induced with methanol,and the expressed product was identified by SDS-PAGE and Western blot.Results The sequence of amplified IL-29 gene was identical to that reported in GenBank(NM_172140).Restriction analysis and sequencing proved that recombinant plasmid pPIC9K-29 was constructed correctly.Specific protein bands with relative molecular masses of about 30 000 and about 27 000 respectively were observed on SDS-PAGE profile of expressed product,both of which showed specific reactions with goat anti-human IL-29 polyclonal antibody.Conclusion Human IL-29 gene was cloned and expressed in P.pastoris GS115,which laid a foundation of further study on biological activity and application of IL-29.