A cloned DNA fragment for identification of Mycobacterium tuberculosis

Abstract

The identification of Mycobacterium tuberculosis is a lengthy process. Attempts were made to develop a more rapid, specific, and sensitive assay for identifying tubercle bacilli in biologic specimens by differentially screening plasmid libraries of Sau3AI-digested M. tuberculosis H37Rv (ATCC 25618) DNA with homologous and heterologous DNA for clones that hybridized strongly with M. tuberculosis DNA alone. Three clones, pMTB1, pMTb2, and pMTb3, were selected for further study. Southern analysis indicated that these clones reacted strongly with DNA from strains of M. tuberculosis isolated in different parts of the world, weakly with DNA from other mycobacterial species, and not at all with Escherichia coli or human DNA. Smaller fragments of mycobacterial DNA contained in plasmid pMTb3 were subcloned into pBR322 (pMTb4) or pUC12 (pMTb5). These recombinant plasmids hybridized with DNA from M. tuberculosis, Mycobacterium bovis, and Mycobacterium bovis BCG (bacille Calmette-Guérin) Montreal and may provide the reagents needed for the development of new methods for rapid diagnosis of M. tuberculosis infections.