FIELD: genetics.
SUBSTANCE: invention relates to genetic engineering. Described is a gene-therapeutic DNA-vector based on a gene-therapeutic DNA-vector VTvaf17, carrying a target gene selected from a group of genes COL1A1, COL1A2, BMP2, BMP7, to increase the level of expression of this target gene in the human body and animals, wherein the gene-therapeutic DNA vector VTvaf17-COL1A1, or VTvaf17-COL1A2, or VTvaf17-BMP2, or VTvaf17-BMP7 has the nucleotide sequence SEQ ID No. 1 or SEQ ID No. 2 or SEQ ID No. 3 or SEQ ID No. 4, respectively. Each of the created genotyping DNA-vectors: VTvaf17-COL1A1, or VTvaf17-COL1A2, or VTvaf17-BMP2, or VTvaf17-BMP7 due to the VTvaf17 vector part limited size, which does not exceed 3,200 base pairs, is capable of efficiently penetrate into cells and to express the target gene cloned therein, selected from the group of COL1A1, COL1A2, BMP2, BMP7 genes. Gene-therapeutic DNA-vector does not contain nucleotide sequences of viral origin and there are no antibiotic resistance genes, providing the possibility of its safe application for genetic therapy of humans and animals. There is also developed a method for obtaining a gene-therapeutic DNA-vector on the basis of gene-therapeutic DNA-vector VTvaf17 carrying the target gene selected from a group of genes: COL1A1, COL1A2, BMP2, BMP7, which consists in that each gene-therapeutic DNA vector: VTvaf17-COL1A1, or VTvaf17-COL1A2, or VTvaf17-BMP2, or VTvaf17-BMP7, are obtained as follows: coding part of the target gene from the group of COL1A1, COL1A2, BMP2, BMP7 genes is cloned into a VTvaf17 DNA-vector and a gene-therapeutic DNA-vector VTvaf17-COL1A1, SEQ ID No. 1 or VTvaf17-COL1A2, SEQ ID No. 2 or VTvaf17-BMP2, SEQ ID No. 3, or VTvaf17-BMP7, SEQ ID No. 4 is obtained, respectively. Method for using the created DNA-vector based on the gene-therapeutic VTvaf17 DNA-vector carrying a target gene selected from a group of genes: COL1A1, COL1A2, BMP2, BMP7, to increase expression level of these target genes consists in introduction of selected gene-therapeutic DNA-vector or several selected gene-therapeutic DNA-vectors into cells, organs and tissues of human or animal, and/or introduction into human and animal organs and tissues of human or animal autologous cells transfected with a selected gene-therapeutic DNA-vector or several selected gene-therapeutic DNA-vectors, or in a combination of said methods. Disclosed is a method of producing Escherichia coli strain SCS110-AF/VTvaf17-COL1A1 or strain Escherichia coli SCS110-AF/VTvaf17-COL1A2, or strain Escherichia coli SCS110-AF/VTvaf17-BMP2, or strain Escherichia coli SCS110-AF/VTvaf17-BMP7 consists in electroporation of competent cells of strain Escherichia coli SCS110-AF with created gene-therapeutic DNA-vector and further selection of stable clones of strain using selective medium. Disclosed is Escherichia coli strain SCS110-AF/VTvaf17-COL1A1, or strain Escherichia coli SCS110-AF/VTvaf17-COL1A2, or strain Escherichia coli SCS110-AFA/Tvaf17-BMP2, or strain Escherichia coli SCS110-AF/VTvaf17-BMP7, carrying a gene-therapeutic DNA-vector for its development with the possibility of culturing the strain without using antibiotics. Method for industrial production of a genotyping DNA-vector consists in scaling a bacterial strain culture to quantities required for growth of bacterial biomass in an industrial fermenter, after which the biomass is used to extract a fraction containing the target DNA product – the gene-therapeutic DNA-vector VTvaf17-COL1A1, or VTvaf17-COL1A2, or VTvaf17-VTvaf17-BMP2, or VTvaf17-BMP7, is multi-step filtered and purified by chromatographic methods.
EFFECT: invention can be used in biotechnology, medicine and agriculture for creation of gene therapy preparations.
14 cl, 14 dwg, 19 ex
