The dissolved oxygen level of any cell Culture environment has a critical effect on cellular metabolism. Specifically, hypoxia condition decreases cell viability and recombinant protein productivity. In this work, to develop CHO cells producing recombinant protein with high productivity, mammalian expression vectors containing a human tissue-type plasminogen activator (t-PA) gene with hypoxia response element (HR-E) were constructed and stably transfected into CHO cells. CHO/2HRE-t-PA cells produced 2-folds higher 2 recombinant t-PA production than CHO/t-PA cells in a Ba2+-alginate immobilized culture, and 16.8-folds in a repeated batch culture. In a non-aerated batch Culture of suspension-adapted cells, t-PA productivity of CHO/2HRE/t-PA cells was 4.2-folds higher than that of CHO/t-PA cells. Our results indicate that HRE is a useful tool for the enhancement of protein productivity in mammalian cell cultures.