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Chapter 17 : Expression of Foreign Genes in Mycobacteria

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Abstract:

This chapter reviews the development of genetic systems for the expression of foreign genes in mycobacteria. Mycobacteriophage have been used for many years to type mycobacterial isolates and have more recently been modified as vectors for efficient delivery of foreign DNA into mycobacteria. Plasmid-based expression systems extend the capabilities of phage-based systems by providing increased cloning capacity, increased copy number in the mycobacterial host, and ease of manipulation. Various derivatives of the pAL5000 plasmid replicon have been combined with plasmid replication origins, such as ColEl and pl5A, by several groups to fashion -mycobacterium shuttle vectors. Expression of foreign proteins in recombinant organisms is influenced by many factors. Much of the increased interest in mycobacteria is a result of the emergence of multidrug-resistant . Complex surface macromolecules, such as oligosaccharides and glycolipids, require the products of many genes and the correct cell envelope structure for proper assembly. BCG, the current vaccine against tuberculosis, has provided variable protective efficacy against tuberculosis in different vaccine trials. Immunization of chickens with recombinant expressing an surface antigen elicits partial protection against coccidiosis.

Citation: Burlein J, Offutt S, Hanson M, Stover C. 1994. Expression of Foreign Genes in Mycobacteria, p 239-252. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch17
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Figure 1

Mycobacterial extrachromosomal (pMV261) and integrative (pMV361) expression vectors (modified from ). Common elements in these two vectors include an expression cassette, the Tn905-derived gene (kanamycin resistance), and an origin of replication functional in derived from pUC19. The expression cassette contains 0.4 kb of the 5ʹ end of the BCG gene, including the promoter region, ribosome-binding site, and first six codons (MAKTIA) as well as a multiple cloning site and the transcriptional terminator. The vectors differ by inclusion of either a mycobacterial plasmid replication origin or the and genes of mycobacteriophage L5.

Citation: Burlein J, Offutt S, Hanson M, Stover C. 1994. Expression of Foreign Genes in Mycobacteria, p 239-252. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch17
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Image of Figure 2
Figure 2

Expression of foreign antigens in different BCG substrains. BCG culture lysates equivalent to approximately 5 x 106 bacteria were analyzed by SDS-PAGE and Coomassie blue staining (A) or by immunoblotting with antibodies specific for foreign antigens OspA (B, C) or HIV-1 gpl20 (D). Panel A shows the total proteins expressed by nonrecombinant BCG. Panels B through D show relevant portions of immunoblots of different BCG recombinants expressing the indicated foreign antigens.

Citation: Burlein J, Offutt S, Hanson M, Stover C. 1994. Expression of Foreign Genes in Mycobacteria, p 239-252. In Bloom B (ed), Tuberculosis. ASM Press, Washington, DC. doi: 10.1128/9781555818357.ch17
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