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Application of CRISPR/Cas9 Genome Editing to Improve Recombinant Protein Production in CHO Cells
  • Grav, Lise Marie ;
  • Rojek, Johan Blatt ;
  • la Cour Karottki, Karen Julie ;
  • Lee, Jae Seong ;
  • Kildegaard, Helene Faustrup
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Publication Year
2025-01-01
Journal
Methods in Molecular Biology
Publisher
Humana Press Inc.
Citation
Methods in Molecular Biology, Vol.2853, pp.49-69
Keyword
Cas9Chinese hamster ovary (CHO) cellsCRISPRGenome editingGlutamine synthetaseKnockoutRecombinant protein production
Mesh Keyword
AnimalsCHO CellsCricetulusCRISPR-Cas SystemsGene EditingGene Knockout TechniquesGlutamate-Ammonia LigaseRecombinant ProteinsRNA, Guide, CRISPR-Cas Systems
All Science Classification Codes (ASJC)
Molecular BiologyGenetics
Abstract
Genome editing has become an important aspect of Chinese hamster ovary (CHO) cell line engineering for improving the production of recombinant protein therapeutics. Currently, the engineering focus is directed toward expanding product diversity while controlling and improving product quality and yields. In this chapter, we present our protocol for using the genome editing tool Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) to knock out engineering target genes in CHO cells. As an example, we describe how to knock out the glutamine synthetase (GS) gene, which increases the selection efficiency of the GS-mediated gene amplification system.
Language
eng
URI
https://aurora.ajou.ac.kr/handle/2018.oak/37045
https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=85207857320&origin=inward
DOI
https://doi.org/10.1007/978-1-0716-4104-0_5
Journal URL
https://www.springer.com/series/7651
Type
Book Chapter
Funding
We thank Karen Kathrine Br\\u00F8ndum and Johnny Arnsdorf for optimizing and setting up FACS sorting and Nachon Charanyanonda Petersen for help with the transfection and FACS sorting. This work was supported by the Novo Nordisk Foundation.
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