Li Qin, Yan Rui-Fang, Yang Yong-Xiao, Mi Chun-Liu, Jia Yan-Long, Wang Tian-Yun
School of Basic Medicine, Xinxiang Medical University, Xinxiang, China.
International Joint Research Laboratory for Recombinant Pharmaceutical Protein Expression System of Henan, Xinxiang Medical University, Xinxiang, China.
Front Bioeng Biotechnol. 2022 May 26;10:840600. doi: 10.3389/fbioe.2022.840600. eCollection 2022.
Chinese hamster ovary (CHO) cells are currently the most widely used host cells for recombinant therapeutic protein (RTP) production. Currently, the RTP yields need to increase further to meet the market needs and reduce costs. In this study, three stabilizing and anti-repressor (SAR) elements from the human genome were selected, including human SAR7, SAR40, and SAR44 elements. SAR elements were cloned upstream of the promoter in the eukaryotic vector, followed by transfection into CHO cells, and were screened under G418 pressure. Flow cytometry was used to detect enhanced green fluorescent protein (eGFP) expression levels. The gene copy numbers and mRNA expression levels were determined through quantitative real-time PCR. Furthermore, the effect of the stronger SAR elements on adalimumab was investigated. The results showed that transgene expression levels in the SAR-containing vectors were higher than that of the control vector, and SAR7 and SAR40 significantly increased and maintained the long-term expression of the transgene in CHO cells. In addition, the transgene expression level increase was related with gene copy numbers and mRNA expression levels. Collectively, SAR elements can enhance the transgene expression and maintain the long-term expression of a transgene in transfected CHO cells, which may be used to increase recombinant protein production in CHO cells.
中国仓鼠卵巢（CHO）细胞是目前重组治疗性蛋白（RTP）生产中使用最广泛的宿主细胞。目前，RTP产量需要进一步提高以满足市场需求并降低成本。在本研究中，从人类基因组中选择了三种稳定和抗阻遏（SAR）元件，包括人类SAR7、SAR40和SAR44元件。将SAR元件克隆到真核载体启动子的上游，然后转染到CHO细胞中，并在G418压力下进行筛选。使用流式细胞术检测增强型绿色荧光蛋白（eGFP）的表达水平。通过定量实时PCR确定基因拷贝数和mRNA表达水平。此外，研究了更强的SAR元件对阿达木单抗的影响。结果表明，含SAR载体中的转基因表达水平高于对照载体，并且SAR7和SAR40显著增加并维持了转基因在CHO细胞中的长期表达。此外，转基因表达水平的提高与基因拷贝数和mRNA表达水平相关。总体而言，SAR元件可以增强转基因表达并维持转基因在转染的CHO细胞中的长期表达，这可用于提高CHO细胞中重组蛋白的产量。
