I-TRAP: a method to identify transcriptional regulator activated promoters.
BACKGROUND
The differential expression of virulence genes is often used by microbial pathogens in adapting to the environment of their host. The differential expression of such sets of genes can be regulated by RNA polymerase sigma factors. Some sigma factors are differentially expressed, which can provide a means to identifying other differentially expressed genes such as those whose expression are controlled by the sigma factor.


METHODS
To identify sigma factor-regulated genes, we developed a method, termed I-TRAP, for the identification of transcriptional regulator activated promoters. The I-TRAP method is based on the fact that some genes will be differentially expressed in the presence and absence of a transcriptional regulator. I-TRAP uses a DNA library in a promoter-trap vector that contains two reporter genes, one to allow the selection of active promoters in the presence of the transcriptional regulator and a second to allow screening for promoter activity in the absence of the transcriptional regulator.


RESULTS
To illustrate the development and use of the I-TRAP approach, the construction of the vectors, host strains, and library necessary to identify SigmaE-regulated genes of Mycobacterium tuberculosis is described.


CONCLUSIONS
The I-TRAP method should be a versatile and useful method for identifying and characterizing promoter activity under a variety of conditions and in response to various regulatory proteins. In our study, we isolated 360 clones that may contain plasmids carrying SigmaE-regulated promoters genes of M. tuberculosis.
McLendon
Molly M
MM
Department of Microbiology and Immunology, Emory University, Atlanta, Georgia 30322, USA. molly-mclendon@uiowa.edu
Shinnick
Thomas M
TM
D008967:Molecular Biology / Q000379:methods*; D009169:Mycobacterium tuberculosis / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000502:physiology*; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic
D012808:Sigma Factor; D014157:Transcription Factors; C063202:sporulation-specific sigma factors