Improvement of the efficiency and quality in developing a new CHO host cell line
- 18 June 2021
- journal article
- research article
- Published by Wiley in Biotechnology Progress
- Vol. 37 (5), e3185
- https://doi.org/10.1002/btpr.3185
Abstract
Chinese hamster ovary (CHO) cells are a ubiquitous tool for industrial therapeutic recombinant protein production. However, consistently generating high-producing clones remains a major challenge during the cell line development process. The glutamine synthetase (GS) and dihydrofolate reductase (DHFR) selection systems are commonly used CHO expression platforms based on controlling the balance of expression between the transgenic and endogenous GS or DHFR genes. Since the expression of the endogenous selection gene in CHO hosts can interfere with selection, generating a corresponding null CHO cell line is required to improve selection stringency, productivity, and stability. However, the efficiency of generating bi-allelic genetic knockouts using conventional protocols is very low (<5%). This significantly affects clone screening efficiency and reduces the chance of identifying robust knockout host cell lines. In this study, we use the GS expression system as an example to improve the genome editing process with zinc finger nucleases (ZFNs), resulting in improved GS-knockout efficiency of up to 46.8%. Furthermore, we demonstrate a process capable of enriching knockout CHO hosts with robust bioprocess traits. This integrated host development process yields a larger number of GS-knockout hosts with desired growth and recombinant protein expression characteristics.Keywords
This publication has 28 references indexed in Scilit:
- Multiplex Genome Engineering Using CRISPR/Cas SystemsScience, 2013
- Improving the efficiency of CHO cell line generation using glutamine synthetase gene knockout cellsBiotechnology & Bioengineering, 2011
- Genome Engineering With Zinc-Finger NucleasesGenetics, 2011
- Gene amplification and vector engineering to achieve rapid and high-level therapeutic protein production using the Dhfr-based CHO cell selection systemBiotechnology Advances, 2010
- Guidelines to cell engineering for monoclonal antibody productionEuropean Journal of Pharmaceutics and Biopharmaceutics, 2010
- Generation of a triple‐gene knockout mammalian cell line using engineered zinc‐finger nucleasesBiotechnology & Bioengineering, 2009
- 25 years of recombinant proteins from reactor-grown cells — Where do we go from here?Biotechnology Advances, 2009
- Design, construction and in vitro testing of zinc finger nucleasesNature Protocols, 2006
- Production of recombinant protein therapeutics in cultivated mammalian cellsNature Biotechnology, 2004
- Deletion of the diploid dihydrofolate reductase locus from cultured mammalian cellsCell, 1983