Identification of expression signals of the mycobacteriophages Bxb1, L1 and TM4 using the Escherichia-Mycobacterium shuttle plasmids pYUB75 and pYUB76 designed to create translational fusions to the lacZ gene

R. G. Barletta, D. D. Kim, S. B. Snapper, B. R. Bloom, W. R. Jacobs

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Abstract

Mycobacterial expression signals were cloned using specially constructed gene fusion shuttle plasmid probes carrying a truncated Escherichia coli lacZ (β-galactosidase) gene which lacked a promoter, a ribosome binding site, and an ATG start codon. Libraries of mycobacteriophage Bxb1, L1 and TM4 DNAs were constructed, and introduced by electroporation into Mycobacterium smegmatis and the 'bacille Calmette-Guerin' (BCG). Clones carrying mycobacterial expression sequences were detected by their blue colour or characteristic fluorescence when plated on media containing chromogenic or fluorogenic substrates. Varying degrees of β-galactosidase expression were observed, and one Bxb1 expression signal was identified where β-galactosidase expression is repressed in phage lysogens.

Original languageEnglish (US)
Pages (from-to)23-30
Number of pages8
JournalJournal of General Microbiology
Volume138
Issue number1
DOIs
Publication statusPublished - Jan 1 1992

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ASJC Scopus subject areas

  • Microbiology

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