| 초록 |
Dissolved oxygen level of cell culture media has a critical effect on cellular metabolism. In this work, to develop CHO cells producing recombinant t-PA with high productivity, mammalian expression vectors containing human tissue-type plasminogen activator (t-PA) gene and hypoxia response element (HRE) were constructed and stably transfected into CHO cells. CHO/2HRE-t-PA cells produced 2 fold higher recombinant t-PA than CHO/t-PA cells in a Ba2+-alginate immobilized culture. The overall t-PA production of immobilized CHO/2HRE/t-PA cells was 334.6 ng ml-1 day-1, total production of 2.68 ug day-1 in a repeated batch culture. It was noted that hypoxia strongly induced CHO cell apoptosis. In this study, bcl-2, an anti-apoptotic gene, was transfected into HRE-dependent or -independent t-PA expressing CHO cells. All cells with bcl-2 gene showed higher t-PA production than those without bcl-2 gene under CoCl2-treated conditions. These results represent the production of recombinant proteins can be increased by introduction of HRE and bcl-2 gene into CHO cells under hypoxia environments. |
