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Effects of different 2A peptides on transgene expression mediated by tricistronic vectors in transfected CHO cells

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Abstract

Multicistronic vectors can increase transgene expression and decrease the imbalance of gene expression in the Chinese hamster ovary (CHO) cell expression system. Small, self-cleaving 2A peptides have a high cleavage efficiency and are essential for constructing high-expression multicistronic vectors. In this study, we investigated the effects of two different 2A peptides on transgene expression in CHO cells via their mediating action on tricistronic vectors. The enhanced green fluorescent protein (eGFP) and red fluorescent protein (RFP) genes were linked by the porcine teschovirus-1 (P2A) and Thosea asigna virus (T2A) peptides in a multicistronic vector. We transfected CHO cells with these vectors and screened for the presence of blasticidin-resistant colonies. Flow cytometry and real-time quantitative PCR (qPCR) were used to detect the expression levels of eGFP and RFP and the copy numbers of stably transfected cells. The results showed that P2A could enhance eGFP and RFP expression by 1.48- and 1.47-fold, respectively, compared to T2A. The expression levels of the genes were not proportional to their copy numbers. In conclusion, we found that P2A can effectively drive transgene expression in CHO cells and a potent 2A peptide can be used for recombinant protein production in the CHO cell system.

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Acknowledgements

This work was supported by the Grants from the National Natural Science Foundation of China (No. 81673337) and the Grants from the Key Scientific Research Projects in Universities of Henan Province (No. 19A350008).

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Xinxiang Medical University, Jinsui Road, Xinxiang, 453003, Henan, People’s Republic of China

    Yan-mei Li, Meng Wang, Tian-yun Wang, Yong-ge Wei, Xiao Guo, Chun-liu Mi, Chun-peng Zhao, Xiang-xiang Cao & Yuan-yuan Dou

  2. International Joint Research Laboratory for Recombiant Pharmaceutical Protein Expression System of Henan, Xinxiang Medical University, Jinsui Road, Xinxiang, 453003, Henan, People’s Republic of China

    Yan-mei Li, Meng Wang, Tian-yun Wang, Yong-ge Wei, Xiao Guo, Chun-liu Mi, Chun-peng Zhao, Xiang-xiang Cao & Yuan-yuan Dou

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  1. Yan-mei Li
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Correspondence to Tian-yun Wang.

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Li, Ym., Wang, M., Wang, Ty. et al. Effects of different 2A peptides on transgene expression mediated by tricistronic vectors in transfected CHO cells. Mol Biol Rep 47, 469–475 (2020). https://doi.org/10.1007/s11033-019-05153-3

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