| DOI | Trouver le DOI : https://doi.org/10.1016/j.jbiotec.2017.06.009 |
|---|
| Auteur | Rechercher : Poulain, A.1; Rechercher : Perret, S.1; Rechercher : Malenfant, F.1; Rechercher : Mullick, A.1; Rechercher : Massie, B.1; Rechercher : Durocher, Y.1 |
|---|
| Affiliation | - Conseil national de recherches du Canada. Thérapeutique en santé humaine
|
|---|
| Format | Texte, Article |
|---|
| Sujet | CHO; cell pools; inducible expression system; methionine sulfoximine; polyethylenimine; recombinant proteins |
|---|
| Résumé | To rapidly produce large amounts of recombinant proteins, the generation of stable Chinese Hamster Ovary (CHO) cell pools represents a useful alternative to large-scale transient gene expression (TGE). We have developed a cell line (CHOBRI/rcTA) allowing the inducible expression of recombinant proteins, based on the cumate gene switch. After the identification of optimal plasmid DNA topology (supercoiled vs linearized plasmid) for PEIpro™ mediated transfection and of optimal conditions for methionine sulfoximine (MSX) selection, we were able to generate CHOBRI/rcTA pools producing high levels of recombinant proteins. Volumetric productivities of up to 900mg/L were reproducibly achieved for a Fc fusion protein and up to 350mg/L for an antibody after 14days post-induction in non-optimized fed-batch cultures. In addition, we show that CHO pool volumetric productivities are not affected by a freeze-thaw cycle or following maintenance in culture for over one month in the presence of MSX. Finally, we demonstrate that volumetric protein production with the CR5 cumate-inducible promoter is three- to four-fold higher than with the human CMV or hybrid EF1α-HTLV constitutive promoters. These results suggest that the cumate-inducible CHOBRI/rcTA stable pool platform is a powerful and robust system for the rapid production of gram amounts of recombinant proteins. |
|---|
| Date de publication | 2017-06-15 |
|---|
| Maison d’édition | Elsevier |
|---|
| Dans | |
|---|
| Langue | anglais |
|---|
| Publications évaluées par des pairs | Oui |
|---|
| Numéro NPARC | 23002175 |
|---|
| Exporter la notice | Exporter en format RIS |
|---|
| Signaler une correction | Signaler une correction (s'ouvre dans un nouvel onglet) |
|---|
| Identificateur de l’enregistrement | ac970221-3075-4921-9c1d-fb59b097f911 |
|---|
| Enregistrement créé | 2017-08-29 |
|---|
| Enregistrement modifié | 2020-03-16 |
|---|
