RECOMBINANT PLASMID DNA PTTG KM2 ENCODING SYNTHESIS OF RECOMBINANT HUMAN IMMUNE INTERFERON, STRAIN ESCHERICHIA COLI T3G - A PRODUCER OF RECOMBINANT HUMAN IMMUNE INTERFERON AND A METHOD OF PREPARING RECOMBINANT HUMAN IMMUNE INTERFERON

摘要

FIELD: molecular biology, genetic engineering. SUBSTANCE: invention relates to construction of recombinant plasmid DNA pTTgKm2 encoding synthesis of human γ--interferon and containing tryptophan promoter from E. coli genome, synthetic sequence of ribosome site binding flanked with restriction sites Eco RI and XbaI, cDNA fragment encoding human γ--interferon, terminator of transcription rrn BTT, gene of resistance to kanamycin (genetic marker) and fragment of vector plasmid pUC19. As a result of transformation of recipient strain E. coli C600 Str^R with recombinant plasmid pTTgKm2 the strain E. coli T3g, a producer of recombinant human γ--interferon, is obtained. Its productivity is up to 1.2 x 1,2×10¹⁰ IU/l cultural fluid. Method of preparing recombinant human γ--interferon based on the use of the constructed recombinant strain E. coli T3g is developed. Method involves the submerged culturing on nutrient medium with decreased tryptophan content, dissolving the aggregated protein in concentrated urea solution with cetavlon, renaturation and purification by treatment with salt of two- or trivalent anions followed by chromatography of solution on anion-exchange resin. EFFECT: enhanced effectiveness of strain, improved method of preparing. 4 cl, 6 dwg, 3 tbl