文献类型：期刊文献

英文题名：φC31-Mediated cassette exchange in Sf9 insect cells for stable expression

作者：Hu, Die[1];Qian, Jingwen[1];Zhang, Tong[1];Yu, Yue[1];Xu, Zhenhe[1];Zhang, Yuanxing[2];Liu, Qin[1,2,3]

机构：[1]East China Univ Sci & Technol, Shanghai Frontiers Sci Ctr Optogenet Tech Cell Met, State Key Lab Bioreactor Engn, Shanghai, Peoples R China;[2]Shanghai Collaborat Innovat Ctr Biomfg, Shanghai, Peoples R China;[3]East China Univ Sci & Technol, Shanghai Frontiers Sci Ctr Optogenet Tech Cell Met, State Key Lab Bioreactor Engn, Shanghai 200237, Peoples R China

年份：2023

卷号：18

期号：7

外文期刊名：BIOTECHNOLOGY JOURNAL

收录：;EI(收录号：20231914062115);WOS:【SCI-EXPANDED(收录号:WOS:000980083500001)】；

基金：Shanghai Municipal Education Commission, Grant/Award Number: 2021Sci & Tech03-28; Fundamental Research Funds for the Central Universities, Grant/Award Number:222201211716

语种：英文

外文关键词：cassette exchange; FACS; insect cell line; Sf9; phi C31

摘要：Insect cells, especially Sf9 cells, are commonly used in biomanufacturing due to their advantages in high expression levels and post-translational modification. However, the development of stable expression cell lines via random integration tended to be unstable. Site-specific integration (SSI) is an alternative strategy. In this study, a fC31 -mediated cassette exchange system in Sf9 cells was established for SSI. The tagging cassette with the reporter gene egfp was randomly inserted into the cell genome. Potential platform cell lines were obtained by fluorescence-activated cell sorting (FACS) and single-cell cloning. Platform cell lines were selected by assessing the fluorescence expression, stability, and growth kinetics of cell lines. The selected platform cell lines were co-transfected with the fC31-containing plasmid and the targeting cassette. Green-fluorescence-negative clones were screened by hygromycin resistance and FACS. The resulting cell clones exhibited the expression properties of the platform cell lines. The rapid development of cell lines for the production of influenza subunit vaccines by the cassette exchange system demonstrated that the system constituted a versatile and reusable platform for the production of various recombinant proteins. Overall, the fC31-mediated cassette exchange system in Sf9 cells has the potential to facilitate and accelerate biologics development.