抄録
Gene amplification has been developed as an indispensable technique for the bio-pharmaceutical production by recombinant mammalian cells. The most commonly used technique is likely dihydroforate reductase (dhfr) gene amplification system in Chinese hamster ovary (CHO) cell line. In this gene amplification system, high productive CHO cells can be selected by "time-consuming" stepwise increases of methotrexiate (MTX) concentration in culture medium. However, it is of great significance to shorten the time for constructing the stable and productive recombinant CHO cell line in the industrial process. In this study, we focused on a rapid construction method for gene-amplified cell line. We employed the human granulocyte-macrophage colony stimulating factor (hGM-CSF) as a model of glycoprotein and investigated the relationship among the productivity and stability of amplified gene, the location of the amplified gene and the MTX concentration. The distribution of amplified gene on the chromosome was analyzed using fluorescence in situ hybridization (FISH). Based on the FISH image analysis, we speculated the specific chromosome location suitable for gene amplification.
