| Literature DB >> 31659692 |
Yan-Mei Li1,2, Meng Wang1,2, Tian-Yun Wang3,4, Yong-Ge Wei1,2, Xiao Guo1,2, Chun-Liu Mi1,2, Chun-Peng Zhao1,2, Xiang-Xiang Cao1,2, Yuan-Yuan Dou1,2.
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.Entities:
Keywords: 2A peptides; CHO cells; Transgene expression; Tricistronic
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Year: 2019 PMID： 31659692 DOI： 10.1007/s11033-019-05153-3
Source DB: PubMed Journal: Mol Biol Rep ISSN： 0301-4851 Impact factor: 2.316