Maximizing the expression of a recombinant gene in Escherichia coli by manipulation of induction time using lactose as inducer.
The use of isopropyl-beta-D-thiogalactoside (IPTG) for induction of the lac-promoter in small-scale cultivations is well established. However, for large-scale microbiological processes the cost of this inducer is a severe limitation. Here is described a method by which lactose is used as inducer of the lac promoter with the same efficiency as that of IPTG. It was found that after growth on glucose the time of the addition of lactose is important for the quality of induction. The resulting yield of the recombinant protein increased when lactose was added to the culture if the glucose concentration was rather low. By careful monitoring of the glucose level in the fermentation, using a biosensor, it was possible to add the inducer when the carbon source was nearly depleted. Using Escherichia coli BL21 (pET3), in which was cloned the main antigen coat protein of the foot and mouth disease virus, induction of the gene led to expression of the target protein at a level exceeding 20% of the total cell protein.
Institut für Angewandte und Technische Mikrobiologie, Ernst-Moritz-Arndt Universität, Greifswald, Federal Republic of Germany.
P
P
Neubauer
K
K
Hofmann
O
O
Holst
B
B
Mattiasson
P
P
Kruschke
D005537:Aphthovirus / Q000235:genetics; D001709:Biotechnology; D002213:Capsid / Q000235:genetics; D036022:Capsid Proteins; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics*; D005814:Genes, Viral / Q000187:drug effects; D007785:Lactose / Q000494:pharmacology*; D010957:Plasmids / Q000187:drug effects; D011994:Recombinant Proteins / Q000096:biosynthesis; D011995:Recombination, Genetic
D036022:Capsid Proteins; D011994:Recombinant Proteins; D007785:Lactose