Kinetics of continuous GM-CSF production by recombinant Saccharomyces cerevisiae in an airlift bioreactor.
Continuous production of murine GM-CSF by recombinant Saccharomyces cerevisiae in an airlift bioreactor was studied at three different dilution rates. The reactor was initially fed with a selective medium to increase cell concentration, and then was fed with a rich, nonselective medium for GM-CSF production. Ethanol was used as the main carbon source to provoke GM-CSF expression. In continuous culture, GM-CSF production was maintained for over 150 h, even though the fraction of plasmid-carrying cells continuously dropped to lower than 20%. The stable GM-CSF production during the later phase of the continuous culture was attributed to increased specific cell productivity possibly resulting from an increase in the plasmid copy number in plasmid-carrying cells. This also indicated the possibility of natural selection of high-copy number cells in continuous culture. Plasmid stability was found to be growth rate (dilution rate) dependent; it increased with the dilution rate. Reactor productivity and specific productivity also increased with the dilution rate. A two-parameter kinetic equation was used to model the plasmid stability kinetics. The growth rate ratio between plasmid-carrying and plasmid-free cells increased from 0.996 to 0.998 while the segregational instability or the probability of plasmid loss from each cell division increased from 1.1% to 16% as the dilution rate decreased from 0.11 h-1 to 0.05 h-1. Oscillation of the dilution rate between 0.05 h-1 and 0.11 h-1 stabilized the plasmids and gave a higher productivity than that achieved without oscillation.
Shu
C H
CH
Department of Chemical Engineering, Ohio State University, Columbus 43210, USA.
Yang
S T
ST
D000818:Animals; D019149:Bioreactors; D001709:Biotechnology; D002452:Cell Count; D000431:Ethanol / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000096:biosynthesis* / Q000235:genetics*; D007700:Kinetics; D051379:Mice; D008957:Models, Genetic; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*
D011994:Recombinant Proteins; D000431:Ethanol; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D005947:Glucose