Effects of viral promoters, the Woodchuck hepatitis post-transcriptional regulatory element, and weakened antibiotic resistance markers on transgene expression in Chinese hamster ovary cells

Abstract

The product productivity of Chinese hamster ovary (CHO) cells need to be further increased and solved. Promoter and selection marker are essential elements for transgene expression. We investigated the effects of promoters, Woodchuck hepatitis post-transcriptional regulatory (WPRE) element, and selection marker on transgene expression in CHO cells. A weakened form of resistance gene neomycin phosphotransferase (NPT) and WPRE element were cloned into the expression vectors driven by the cytomegalovirus (CMV) and simian virus 40 (SV40) promoters, following by transfecting into CHO cells and screened under G418 pressure. qRT-PCR and Flow cytometry were used to detect the expression level of eGFP gene, and transgene copy number was determined using qPCR technology. Western blot and ELISA were utilized to analyze the expression levels of cetuximab. The weakened NPT gene enhanced eGFP expression both under the CMV or SV40 promoters. Moreover, WPRE, combined with the NPT gene considerably enhanced eGFP expression in CHO cells. Furthermore, the CMV promoter and WPRE combination resulted in higher cetuximab expression levels than CMV promoter alone. However, WPRE could not enhance the transgene expression under the circumstance of CMV promoter and weakened NPT gene combination, suggesting that the effects of the WPRE combined with selection markers were specificity.