Biopolym. Cell. 2003; 19(5):457-462.
Molecular and Cell Biotechnologies
Influence of various inductor concentrations on the human alpha-2b interferon production in the bacteriophage T7 RNA polymerase-base expression system in Escherichia coli cells
1, 2Slavchenko I. Yu., 1Boreyko E. V., 1Vorobey N. V.
  1. Institute of Molecular Biology and Genetics, NAS of Ukraine
    150, Akademika Zabolotnoho Str., Kyiv, Ukraine, 03680
  2. PSRS "Biotechnolog"
    150, Akademika Zabolotnogo Str., Kyiv, Ukraine, 03680

Abstract

Various concentrations (from 0.01 up to JO MM) of isopropyl-fi-D-thiogalactoside (IPTG) were used to induce the human atpha~2b interferon (IFN) biosynthesis by recombinant E. coli BL21 (DE3) cells, harboring plasmid pET-alFN. The plasmid pET-alFN carries an artificial gene for IFN, which is under the control of the gene 10 phage T7 promoter. The gene for phage T7 RNA polymerasc is situated in the chromosome of E. coli BL2I (DE3) cells under the control of the lac-UV5 E. coli promoter. We have found, that a minimum concentration of IPTG, ensuring effective expression of the target protein, increases with the lowering of the producer cultivation temperature after induction. Thus, the full induction is achieved by adding IPTG to the medium in final concentration 0.06 mM at 37 °C, and 0,1 mM at 21 °C. It was shown, that the decrease of inductor concentration up to 0.01 mM did not promote accumulation of IFN in a soluble form when producer was cultivated at the temperature of 37 °C. It was also determined that IPTG in final concentration up to 2 mM did not have negative effect on the lytic development of bacteriophage lambda. As a result of the lysis of E. coli BL (pET-alFN) cells, infected by phage, the recombinant protein in a soluble form accumulates immediately in a growth medium. The minimum concentration of IPTG, ensuring the maximum yield of IFN obtained by this method is 0.2 mM.

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