1887

Abstract

An –mycobacterial shuttle vector, pJCluc, containing a luciferase reporter gene, was constructed and used to analyse the promoter. A 19 kb region immediately upstream of promoted expression of the luciferase gene in and . A smaller promoter fragment (559 bp) promoted expression with equal efficiency, and was used in all further studies. Two transcription start sites were mapped by primer extension analysis to 47 and 56 bp upstream of the GTG initiation codon. Putative promoters associated with these show similarity to previously identified mycobacterial promoters. Deletions in the promoter fragment, introduced with BAL-31 nuclease and restriction endonucleases, revealed that a region between 559 and 448 bp upstream of the translation initiation codon, designated the upstream activator region (UAR), is essential for promoter activity in , and is required for optimal activity in . The UAR was also able to increase expression from the promoter 15-fold in and 12-fold in . An alternative promoter is active in deletion constructs in which either the UAR or the promoters identified here are absent. Expression from the promoter peaks during late exponential phase, and declines during stationary phase. The promoter is induced by ascorbic acid, and is repressed by oxygen limitation and growth at elevated temperatures. The promoter constructs exhibited similar activities in BCG as they did in .

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1999-09-01
2019-12-13
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