Efficient production of recombinant proteins in suspension CHO cells culture using the Tol2 transposon system coupled with cycloheximide resistance selection

Abstract

DNA recombination techniques in mammalian cells has been applied to the production of therapeutic proteins for several decades. To be used for commercial production, established cell lines should stably express target proteins with high productivity and acceptable quality for human use. In the conventional transfection method, the screening process is laborious and time-consuming since superior cell lines had to be selected from an enormous number of transfected cell pools and clonal cell lines with a wide variety of transgene insertion locations. In this study, we demonstrated that the combination of a Tol2 transposon system and cell selection by cycloheximide resistance is an efficient method to express therapeutic proteins, such as human antibody in suspension culture of Chinese hamster ovary cells. The resulting stable cell lines showed constant productivity and cell growth over a long enough cultivation periods for recombinant protein production. We anticipate that this approach will prove widely applicable to protein production in research and development of pharmaceutical products.

Promoting the production of challenging proteins via induced expression in CHO cells and modified cell-free lysates harboring T7 RNA polymerase and mutant eIF2α

“…Production of biotechnologically relevant proteins can be achieved by expression of the protein of interest in cultured cells or using cell-free reaction conditions. In cell-based approaches, transient as well as stable transfection is often used for recombinant protein production [ 1 , 2 ]. On the other hand, protein synthesis in cell-free systems can be performed by adding DNA templates to the open reaction, which is driven by the presence of an active cell lysate containing components for transcription and protein translation including ribosomes, aminoacyl tRNA synthetases (aaRS), transcription and translation factors, viral RNA polymerase for transcription, substrates and an energy regeneration system [ 3 , 4 ].…”