Effects of viral promoters,the Woodchuck hepatitis post-transcriptional regulatory element,and weakened antibiotic resistance markers on transgene expression in Chinese hamster ovary cells |
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| Affiliation: | 1. Department of Biochemistry and Molecular Biology, Xinxiang Medical University, Xinxiang, 453003, Henan, China;2. International Joint Research Laboratory for Recombinant Pharmaceutical Protein Expression System of Henan, Xinxiang Medical University, Xinxiang, Henan, China;1. Cancer Biology Laboratory, Department of Biochemistry, School of Life Sciences, Bharathidasan University, Tiruchirappalli, 620 024, Tamil Nadu, India;2. Department of Animal Science, Chonbuk National University, Jeonju, 561-756, Republic of Korea;3. Crystal Growth and Thin Film Laboratory, Department of Physics, Bharathidasan University, Tiruchirappalli, 620024, Tamil Nadu, India;1. Biorefinery and Bioprocess Engineering Laboratory, Department of Chemical Engineering and Materials Science, Yuan Ze University, Chungli, Taoyuan, 320, Taiwan;2. Faculty of Applied Sciences, UCSI University, UCSI Heights, 56000 Cheras, Kuala Lumpur, Malaysia;1. Fraunhofer Institute of Interfacial and Bioprocess Engineering, Stuttgart, Germany;2. Fraunhofer Center for Chemical-Biotechnological Processes, Leuna, Germany;1. Processes Engineering Laboratory, State University of Bahia Southwest, Itapetinga, Bahia, Brazil;2. Department of Animal and Rural Technology, State University of Southwest of Bahia, 45700-000 Itapetinga, Bahia, Brazil;3. Department of Exact and Natural Sciences, State University of Southwest of Bahia, 45700-000 Itapetinga, Bahia, Brazil;4. Department of Natural Science, State University of Southwest of Bahia, 45083-900 Vitoria da Conquista, Bahia, Brazil;1. State Key Laboratory of Biochemical Engineering, Beijing Key Laboratory of Biomass Refining Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, PR China;2. University of Chinese Academy of Sciences, Beijing 100049, PR China;1. Key Laboratory of Food Bioengineering (China National Light Industry), College of Food Science and Nutritional Engineering, China Agricultural University, Beijing 100083, China;2. Beijing Advanced Innovation Center for Food Nutrition and Human Health, College of Engineering, China Agricultural University, Beijing 100083, China |
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| 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. |
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| Keywords: | Chinese hamster ovary WPRE Promoter Weakened resistance gene Transgene |
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