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Cloning, characterization and disruption of a (p)ppGpp synthetase gene ( relA ) of Streptomyces coelicolor A3(2)
Author(s) -
Chakraburtty Rekha,
White Janet,
Takano Eriko,
Bibb Mervyn
Publication year - 1996
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1046/j.1365-2958.1996.390919.x
Subject(s) - streptomyces coelicolor , biology , cloning (programming) , gene , genetics , molecular cloning , gene expression , mutant , computer science , programming language
An internal segment of the (p)ppGpp synthetase gene, relA , of Streptomyces coelicolor A3(2) was amplified from genomic DNA using the polymerase chain reaction and used as a hybridization probe to isolate the complete gene from a cosmid library. relA lies downstream of a gene ( apt ) that apparently encodes adenine phosphoribosyltransferase and is transcribed from two promoters, relA p1 and relA p2, and by transcriptional readthrough from apt . While the level of relA p2 transcripts remained relatively constant, relA p1 activity apparently peaked during transition phase, following a decline in readthrough transcription from apt . Disruption of relA using an att − derivative of the temperate phage φC31 abolished ppGpp synthesis on amino acid depletion. When grown on agar, the disruptants grew more slowly than a control lysogen made with an att + phage vector and gave smaller colonies that sporulated normally. The relA mutation had no consistent or marked effect on actinorhodin production in either liquid‐ or agar‐grown cultures, indicating that elevated levels of (p)ppGpp are not essential for triggering the onset of antibiotic production.