RECOMBINANT PLASMID DNA PERPINS1 ENCODING AMINO ACID SEQUENCE OF HUMAN RECOMBINANT PROINSULIN AND STRAIN ESCHERICHIA COLI BL21(DE3)/PERPINS1 AS PRODUCER OF HUMAN RECOMBINANT PROINSULIN

摘要

FIELD: biotechnology, genetic and protein engineering, molecular biology. SUBSTANCE: invention relates to preparing high-productive recombinant bacterial strain-producer of human insulin. For this purpose the strain-recipient is transformed with recombinant plasmid DNA pERPIns1 comprising of DNA fragment Nde I/Hind III of plasmid pET 20b(+) including promoter and terminator of transcription of T7- RNA polymerase, enhancer of translation of gene 0 of phage T7, β-lactamase gene and DNA fragment Nde I/Hind III containing gene sequence of recombinant human proinsulin adapted to these sites. Strain-producer is cultured up to accumulation human recombinant proinsulin in the amount 60-70% of total cellular protein. Cells are disrupted in buffer solution and inclusion bodies are isolated. Strain-producer E. coli BL21 (DE3)/pERPIns is grown in rich medium (YT-, LB-broth and others) (or induced with isopropylthio-β-D- galactoside and grown again) up to maximal density value of culture. Invention provides preparing human recombinant proinsulin. EFFECT: simplified technology, high yield of proinsulin. 2 cl, 1 dwg, 2 ex