METHOD OF PRODUCING RECOMBINANT VP3 VIRUS OF INFECTIOUS BURSAL DISEASE Russian patent published in 2019 - IPC C12N15/34 C12N15/64 C12N15/85 

FIELD: medicine.

SUBSTANCE: disclosed is a method of producing recombinant VP3 protein of infectious bursal disease virus, involving creation of a genetic engineering structure. Human embryonic kidney cells (line Expi293F) are used, transfection of which is carried out with a plasmid pEG-VP3-His containing an insert on the HindIII and BamHI restriction sites of the nucleotide sequence coding the mature VP3 protein of the DDI IBDV strain with 6-histidine portion at the C-terminal and the stop codon (nucleotide sequence of the gene VP3-His – SEQ ID No. 1), under immediate-early promoter control of cytomegalovirus, as well as containing a gene providing resistance to neomycin and kanamycin, under the control of the early SV40 virus gene promoter. Clones growing on selective marker G-418 are selected. Clones are analyzed for recombinant VP3 production by means of western blot technique with monoclonal antibodies on histidine tag. Cells of the target clone are transformed into a suspension culture, cultured, and a cytoplasmic fraction of the cells of the line Expi293F/VP3-His is obtained. Recombinant VP3 protein is extracted from said cytoplasmic cell fraction by affinity chromatography on a Ni-NTA sepharose sorbent column, followed by analysis of antigenic activity of recovered recombinant VP3 protein.

EFFECT: invention provides producing stable human cell lines producing properly folded and highly immunogenic VP3 protein.

1 cl, 6 dwg, 6 ex