Abstract
The generation of the stable, high-level recombinant protein-producing cell lines remains a significant challenge in the biopharmaceutical industry. Expression vector optimization is an effective strategy to increase transgene expression levels and stability, and the choice of suitable poly A element is crucial for the expression of recombinant protein. In this study, we investigated the effects of different poly A elements on transgene expression in Chinese hamster ovary (CHO) cells. Five poly A elements, including bovine growth hormone (BGH), mutant BGH, herpes simplex virus type 1 thymidine kinase (HSV-TK), SV40, and a synthetic (Synt) poly A, were cloned into the expression vector and transfected into CHO cells. The results indicated the SV40 and Synt poly A sequences can significant improve eGFP transgene expression in stable transfected CHO cells and maintain long-term expression. However, qPCR results showed that the eGFP expression at protein level was not related to the gene copy number and mRNA level. Importantly, the SV40 and Synt poly A elements decreased the variation of eGFP transgene expression. Furthermore, it also showed that the SV40 and Synt poly A elements induced higher levels of adalimumab expression. In conclusion, SV40 poly A and Synt poly A are stronger elements that increase stable transgene expression and decrease the variation of expression, and the choice of suitable poly A element is helpful to improve the expression of recombinant protein.
Highlights
Chinese hamster ovary (CHO) cells are the most frequently used expression system for the production of recombinant therapeutic proteins (Tripathi and Shrivastava 2019; Kuo et al, 2018)
We investigated the effects of different poly A elements, including bovine growth hormone (BGH), M BGH, Synt, herpes simplex virus type thymidine kinase (HSV-TK), and SV40 poly A, on transgene expression and variation of expression in transfected CHO cells
The results indicated that polyadenylation sequences can influence transgene expression, different poly A showed different transgene expression levels, and the SV40 and Synt poly A significantly increased transgene expression levels and stability, decreased variation of expression
Summary
Chinese hamster ovary (CHO) cells are the most frequently used expression system for the production of recombinant therapeutic proteins (Tripathi and Shrivastava 2019; Kuo et al, 2018). High recombinant protein production, cell line engineering, growth medium modification, and the incorporation of cisacting elements were performed, and obtained the ideal purpose (Jia et al, 2018; Guo et al, 2020). Among these approaches, expression vector optimization is an effective strategy to increase transgene expression level and stability. Most studies have focused on the role of the enhancer, internal ribosome entry site (IRES), intron, promoters, and combinations of regulatory elements with promoters (Hunter et al, 2019; Skipper et al, 2019; Yeo et al, 2017; Chai et al, 2018), few studies have been performed on the effects of poly A elements on transgene expression in CHO cells
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