Identification of strong promoter elements of Mycobacterium smegmatis and their utility for foreign gene expression in mycobacteria

• Published in: FEMS microbiology letters Vol. 224; no. 1; pp. 139 - 142
• Main Authors: Spratt, Joanne M, Britton, Warwick J, Triccas, James A
• Format: Journal Article
• Language: English
• Published: Oxford, UK Elsevier B.V 15.07.2003; Blackwell Publishing Ltd; Blackwell
• Subjects: Antigens, Bacterial - genetics; Bacteriology; Biological and medical sciences; Fundamental and applied biological sciences. Psychology; Gene expression; Gene Expression Regulation, Bacterial; Genetics; Green fluorescent protein; Green Fluorescent Proteins; Indicators and Reagents - metabolism; Luminescent Proteins - genetics; Microbiology; Molecular and cellular biology; Molecular genetics; Mycobacterium leprae - genetics; Mycobacterium smegmatis; Mycobacterium smegmatis - genetics; Promoter Regions, Genetic - genetics; Protein purification; Strong promoter; Vaccines, antisera, therapeutical immunoglobulins and monoclonal antibodies; Mycobacterium smegmatis; Strong promoter; Protein purification; Green fluorescent protein; Characterization; Promoter; Purification; Mycobacteriales; Mycobacteriaceae; Bacteria; Actinomycetes; Gene expression; Comparative study; Protein
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1016/S0378-1097(03)00442-7

The isolation of elements driving high-level expression of foreign genes in mycobacteria would significantly aid characterization of mycobacterial antigens and recombinant vaccine development. Mycobacterium smegmatis is a widely employed host for recombinant mycobacterial gene expression. This report describes the identification of strong promoter elements of M. smegmatis. Fluorescence-activated cell sorting was employed to isolate DNA fragments permitting high-level expression of the Aequorea victoria green fluorescent protein within recombinant M. smegmatis. Ten postulated M. smegmatis promoters were identified which showed activity two to six times that of the strong β-lactamase promoter of Mycobacterium fortuitum. The utility of one of these promoters for the over-expression of foreign genes in mycobacteria was demonstrated by the efficient purification of the Mycobacterium leprae 35-kDa antigen from recombinant M. smegmatis.