PEI-Mediated Transient Gene Expression in CHO Cells

• Published in: Methods in molecular biology (Clifton, N.J.) Vol. 1850; p. 33
• Main Author: Rajendra, Yashas
• Format: Journal Article
• Language: English
• Published: United States 01.01.2018
• Subjects: Animals; CHO Cells; Cricetulus; Polyethyleneimine - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Transfection - methods; Polyethyleneimine; Transfection; Recombinant protein; CHO cells; Orbital shaking
• DOI: 10.1007/978-1-4939-8730-6_3

We describe a method for polyethyleneimine (PEI) mediated transient transfection of suspension-adapted Chinese hamster ovary (CHO-DG44) cells for protein expression applicable at scales from 2 mL to 2 L. The method involves transfection at a high cell density (5 × 10 cells/mL) by direct addition of plasmid DNA (pDNA) and PEI to the culture and subsequent incubation at 31 °C with agitation by orbital shaking. This method requires 0.3 mg/L of coding pDNA, 2.7 mg/L of nonspecific (filler) DNA and 15 mg/L of PEI. The production phase is performed at 31 °C in the presence of 0.25% N,N-dimethylacetamide (DMA). We also provide information on culture vessel options, recommended working volumes, and recommended shaking speeds for transfections at scales from 2 mL to 2 L.