Construction Of CHO Cell Line Stably Expressing Mouse Lamada-Interferon

Objective The Construction of the eukaryotic expressing vector pEGFP-mIFN-λ2 and the followed transfectinon of the recombinant into CHO cells aim at obtaining the CHO cell line which expresses the IFN-)λ2 of mouse stably.The biological activity of mIFN-λ2 which expressed by the CHO-mIFN-λ2 cell line was studied at the cells cultured in vitro.This study lays not only a foundation for a further study of the biological function of the IFN-λs but also for a study of the impact of the IFN-λon lung cancer in vivo.Methods Full-length of mIFN-λ2 cDNA was obtained by using RT-PCR from cells of mouse spleen stimulated by vesicular stomatovirus and then subcloned to PMD18-T Vector for sequencing.The confirmed gene mIFN-λ2 was then subcloned to the eukaryotic expressing vector pEGFP.CHO cells were transfected stably by the eukaryotic expressing vector pEGFP- mIFN-λand G418 was used for obtaining postive clone cells.The product of postive clone cells was used for studying the antivirat activity of the mIFN-λ2 in the cell line of lung cancer.Results The recombinant PMD18-T-mIFN-λ2 was cut by two kinds of enzyme,SacⅠand XhoⅠ,the fragment was 582 bp,and the analysis of sequence shows it was entirely consistent with nucleotide sequences reported in GenBank.pEGFP-mIFN-λ2 eukaryotic expressing vector was constructed successfully and expressed stably in CHO cells,and the mRNA of mIFN-λ2 was verified expressing in CHO-pEGFP-mIFN-λ2 cell line by RT-PCR.The antivirus activity of the supernatant secreted by the CHO- mIFN-λ2 cell line was 10~4 AU/ml.Conclusions Gene cloning of mIFN-λ2 was successful.The eukaryotic expressing vector of mIFN-λ2 was constructed successfully and stably in CHO cells,and its product has obvious antivirus activity in vitro.It is then proved that the mIFN-λ2 protein we have expressed has a relatively high biological activity.And it lays not only a solid foundation for a further study of the biologic function of the IFN-λs but also for a study of the impact of the IFN-λon lung cancer in vivo.