METHOD FOR IDENTIFYING NEW TARGET FOR USING IN DEVELOPMENT OF THERAPEUTIC FORMULATIONS AND MEDICINAL AGENTS SHOWING EFFECTIVENESS AGAINST TUBERCULOSIS

摘要

FIELD: biotechnology, medicine, pharmaceutical industry, genetic engineering.;SUBSTANCE: method involves the following steps: destruction of gene devR in plasmid pJT53.34, construction of plasmid pJQ2000KdevR : kan from the destructed gene and its insertion into M. tuberculosis H37Rv cells followed by selection of transformants in Middlebruck medium 7H10 with kanamycin. Then transformants are checked for the presence of sequences of genes devR, Km^R and gene determining resistance to sucrose (SacB) and M. tuberculosis Dup devR comprising copy of wild type and destructed copy of locus devR are detected. After carried out analysis these mycobacteria are grown in the same medium in the presence of kanamycin and sucrose and the following multiplication of mycobacteria under the same culturing conditions. The strain M. tuberculosis with mutation with respect to devR is checked for confirming allel change, destruction of gene devR : kan, among them and its functional destruction, and its growth properties are evaluated also. Then analysis for the presence of transcripts obtained from operon Rv 3134c - devR - devS is carried out and also their scanning and testing the virulence of the obtained mutant by devR strain in guinea pigs. Analysis involves histopathological analysis of infected organs of guinea pigs, isolation of identified mutant mycobacteria from spleens of infected animals and quantitative analysis of bacterial charge. Use of invention allows carrying out the identification of both a target responsible for relapse/reactivation of this diseases in patient and a target that allows adaptation of organism to hypoxia, and, respectively, to develop therapeutic formulations and medicinal agents for effective treatment of tuberculosis.;EFFECT: improved identifying method of target.;6 cl, 4 dwg