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PMID: 384366 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S. Research Support, U.S. Gov't, P.H.S.

Characterizing wild-type and mutant promoters of the tetracycline resistance gene in pBR313.

Nucleic acids research ·Vol. 6 ·No. 10 ·1979-07-25 ·Pages 3267-87

Rodriguez RL, West RW, Heyneker HL, Bolivar F, Boyer HW

Abstract

By employing a system of RNA polymerase binding and restriction endonuclease digestion, we demonstrate that the region in and around the Hind III site of pBR313 and pBR322 is the promoter for the tetracycline (Tc) resistance gene(s). Furthermore, it is shown that this region was transferred intact from pSC101 during the construction of the latter plasmids. The in vitro insertion of a few base pairs at the Hind III site produces a series of "down" promoter mutations in which the level of in vivo Tc resistance is reduced. Sequence analysis of the various promoter mutations revealed significant base pair rearrangements in the region between -40 and -12 of the promoter. While these base alterations do not appear to affect the firm binding of RNA polymerase, they do affect the ability of mutant promoters to initiate transcription. These observations suggest that the region from -40 to -12, previously designated as the "recognition region", is actually involved in the process of initiation of transcription.

MeSH Terms
Base Sequence DNA Restriction Enzymes DNA-Directed RNA Polymerases/metabolism Drug Resistance, Microbial Escherichia coli/metabolism Mutation/drug effects Plasmids Tetracycline/pharmacology
Chemicals
DNA-Directed RNA Polymerases DNA Restriction Enzymes Tetracycline
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Rodriguez R L
West R W
Heyneker H L
Bolivar F
Boyer H W
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38 references, click to expand
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Article Info
Journal
Nucleic acids research
Abbr.
Nucleic Acids Res
ISSN
0305-1048
DOI
10.1093/nar/6.10.3267
Published
1979-07-25
Pages
3267-87
Language
English
Region
England
NLM ID
0411011
PMCID
PMC327933
Subset
IM
External Links
PubMed source DOI: 10.1093/nar/6.10.3267 PMC Full Text View journal details
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