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

by Selleckchem | May 16 2023

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.

Comments:

The study demonstrates an efficient method for producing therapeutic proteins using DNA recombination techniques in mammalian cells. The method involves combining the Tol2 transposon system with cell selection by cycloheximide resistance. The resulting stable cell lines express the target proteins with high productivity and acceptable quality for human use.

The conventional transfection method for producing stable cell lines is laborious and time-consuming. The screening process involves selecting superior cell lines from a large pool of transfected cells and clonal cell lines with a wide variety of transgene insertion locations. The method proposed in this study reduces the time and effort required for the screening process, making it more efficient.

The Tol2 transposon system is a well-established method for introducing foreign DNA into the genome of mammalian cells. The system consists of a transposon vector containing the gene of interest flanked by Tol2 transposase recognition sequences. The transposon vector is co-transfected with a plasmid encoding the Tol2 transposase enzyme into the target cells. The transposase enzyme mediates the transposition of the transposon vector into the genome of the target cells, resulting in stable integration of the transgene.

Cycloheximide is an antibiotic that inhibits protein synthesis by binding to the ribosome. The use of cycloheximide selection allows for the isolation of cells that have integrated the transgene and are expressing the protein of interest. The resulting stable cell lines exhibit constant productivity and cell growth over long cultivation periods, making them suitable for the commercial production of recombinant proteins.

In summary, the combination of the Tol2 transposon system and cell selection by cycloheximide resistance provides an efficient method for producing stable cell lines that express therapeutic proteins with high productivity and acceptable quality for human use. This method has potential applications in research and development of pharmaceutical products.