Double knock-out CHO cell line method of its generation and producing therapeutic proteins therefrom

Abstract

The present disclosure relates to providing an engineered CHO cell line wherein the two essential metabolic genes are knocked out. Particularly, the present invention relates to a double knockout CHO cell line (DHFR−/− and GS−/−) with disrupted Dihydrofolate Reductase (DHFR) and Glutamine Synthetase (GS) genomic loci. The double knockout CHO cell line (DHFR−/− and GS−/−) being suitable for expression of monoclonal antibodies, dimeric therapeutic proteins, Fab, single chain fragments, or the like. The present disclosure also provides method of generation of a double knock out CHO cell line (DHFR−/− and GS−/−) using gene selection and/or manipulating techniques such as CRISPR/Cas9 system, Zinc Finger Nuclease, TALEN, or the like. The present disclosure further provides method of selection of clones and production of therapeutic proteins of interest with increased titre.

Dependencies are inferred from the claim text.