Establishment of a cryopreservation method for the industrial use of D-amino acid oxidase-overexpressing Escherichia coli.
A cryopreservation condition for D-amino acid oxidase (DAAO)-overexpressing Escherichia coli (E. coli BL21(DE3)/pET-DAAO) was established. Ten percent was the optimum concentration of glycerol as a cryoprotectant, and its diffusion into stationary phase cells was superior to that into log cells. The results also showed that rather than fast cooling, a slow cooling method was appropriate to our recombinant E. coli. In addition, 15 min was the best equilibration period, at which higher than 90% of recovery rates were maintained at all test points. Most importantly, the relative recovery rates, product yield, and fermentation pattern of the cell banks (CBs) constructed according to our cryopreservation method did not change over 12 months, confirming that our method not only permits exceptional cryopreservation, but offers prolonged productivity. Taken together, our results demonstrating a cryopreservation method for E. coli BL21(DE3)/pET-DAAO provide insight into an improvement in the industrial production of DAAO.
Kim
Sae-Jin
SJ
School of Life Sciences and Biotechnology, Korea University, Seoul.
Park
Hye Won
HW
Shin
Chang-Hun
CH
Kim
Chan-Wha
CW
D001709:Biotechnology / Q000379:methods*; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents / Q000494:pharmacology; D003605:D-Amino-Acid Oxidase / Q000096:biosynthesis* / Q000235:genetics*; D004926:Escherichia coli / Q000166:cytology* / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D005285:Fermentation; D015870:Gene Expression; D005818:Genetic Engineering; D005990:Glycerol / Q000494:pharmacology; D013997:Time Factors
D003451:Cryoprotective Agents; D003605:D-Amino-Acid Oxidase; D005990:Glycerol