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Published on: December 19, 2011
Enhanced transgene expression using cis-acting elements combined with the EF1 promoter in a mammalian expression
Wen Wang1, Xiao Guo2, Yan-Mei Li3
1Department of Biochemistry and Molecular Biology, Xinxiang Medical University, Xinxiang 453003, Henan, China; International Joint Research Laboratory for Recombinant Pharmaceutical Protein Expression System of Henan, Xinxiang Medical University, Xinxiang, Henan, China; College of Pharmacy, Xinxiang Medical University, Xinxiang 453003, Henan, China.
Abstract:
Transgene expression in eukaryotic cells suffers from epigenetic effects that result in low or unstable transgene expression and high clonal variability. The use of epigenetic regulators is a promising approach to alleviating such unwanted effects. we investigated the effect of the strong human EF1-α promoter combined with six cis-acting elements on transgene expression in transfected Chinese hamster ovary (CHO) cells. The six elements included the human cytomegalovirus immediate early core promoter element (hCPE), three synthetic enhancer element (SEE1, SEE2, Syn1), human cytomegalovirus immediate early enhancer (hCMV-IEE), and a regulatory element isolated from CHO-K1 genomic DNA (C77). The respective vectors were transfected into CHO cells, and stably transfected cell pools were screened and analyzed for transgene expression. The results showed that SEE1 increased transient expression most strongly. However, hCPE enhanced eGFP transgene expression most significantly in stably transfected CHO cells, by about 2.45-fold. Erythropoietin expression analysis showed that hCPE induced the highest EPO productivity, followed by hCMV-IEE. We found that the enhancing effect of hCPE and hCMV-IEE was related with transgene copy number. In conclusion, we found that hCPE and hCMV-IEE cis-acting elements combine with EF1-α can increase recombinant protein expression in CHO cells.
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