METHOD OF PRODUCING RECOMBINANT ANTIMICROBIAL PEPTIDE UBI18-35, RECOMBINANT PLASMID DNA pET31b-2xUBI18-35 AND PRODUCER-PRODUCER OF ESCHERICHIA COLI BL21 ROSETTA DE3 pLusS/pET31b-2HUBI18-35 ANTIMICROBIAL PEPTIDE UBI18-35 Russian patent published in 2019 - IPC C12N15/70 C12N1/21 C12N15/12 C12R1/19 

FIELD: biotechnology.

SUBSTANCE: invention relates to biotechnology and represents recombinant plasmid DNA pET31b-2xUBI18-35 coding amino acid sequence of recombinant antimicrobial peptide UBI18-35 coded by nucleotide sequence: 5’-AAAGTGGCGAAACAGGAAAAGAAAAAGAA AAAGACCGGTCGTGCGAAACGTCGT-3’, having molar mass of 1.8 MPa. Plasmid DNA consists of DNA fragment of plasmid pET31b + hydrolysed at AlwNI (PstNI) site ligated on said site with fragment coding 2 UBI18-35 molecules bounded from both ends with trinucleotides, coding methionine residues containing a plasmid pBR322 replication initiation site, a T7-RNA polymerase phage transcription promoter and terminator, a starter codon, a coding gene for ketosteroidisomerase, a coding sequence of 6 histidine residues, a stop codon, a genetic marker AMPr - a β-lactamase gene which determines the resistance of the transformed plasmid pET31b-2xUBI18-35 cells Escherichia coli to ampicillin. Invention also relates to a method of producing recombinant antimicrobial peptide UBI18-35, comprising cell transformation Escherichia coli BL21 Rosetta DE3 pLysS by said plasmid, growing cells of transformed cells Escherichia coli in medium with subsequent cultivation; separation of cells by centrifugation, washing of inclusion bodies containing 2 molecules of recombinant antimicrobial peptide UBI18-35 in protein-fusion with ketosteroidisomerase and 6-histidine tag, dissolving them in a buffer with addition of urea, followed by separation of peptides from ketosteroidisomerase and 6 histidine residues, separation thereof using bromocyan and chromatographic purification of the recombinant peptide.

EFFECT: plasmid DNA provides synthesis of UBI18-35 peptide as a part of fusion protein with ketosteroidisomerase.

2 cl, 9 dwg, 1 tbl