Abstract
Genetic chracterstics of the structural gene of guamerin (a novel elastase inhibitor from Korean leech), integrated into the HIS4 locus of chromosomal DNA of Pichia pastoris along with the $\alpha$-factor leader sequence, were investigated. In the selected clone from candidates, two copies of the integration cassette including the structural gene copies of the integration cassette including the structural gene of guamerin were found in the integration site of the chromosomal DNA of P.pastoris. It was demonstrated that the integrated structural gene of guamerin was stable up to about 70 generations in the relay flask culture. Then, a high-cell-density culture could be fulfilled easily by DO-stat fed-batch culture, in which the cell growth and the recombinant guamerin production reached about 250 of OD600nm and 260 mg/l, respectively. Finally, it was revealed that the DNA sequence of the integrated structural gene of guamerin in P. pastoris was maintained correctly in the end of production cells of relay flask culture and high-cell-density culture.
