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Nucleotide sequence of an Rts1 fragment causing temperature-dependent instability.
1Department of Microbiology, School of Medicine, University of Pennsylvania, Philadelphia 19104-6076.
Journal of Bacteriology
|March 1, 1988
Summary
The Rts1 plasmid exhibits temperature-dependent instability (Tdi), leading to its elimination from bacteria at higher temperatures. This Tdi phenotype is controlled by a specific DNA segment, requiring a particular open reading frame for its expression.
Area of Science:
- Molecular Biology
- Genetics
- Microbiology
Background:
- The Rts1 plasmid is known for its multiphenotypic drug resistance.
- It displays temperature-dependent instability (Tdi), being eliminated from host bacteria at 42°C but not at 32°C.
Purpose of the Study:
- To determine the nucleotide sequence of the Rts1 DNA b' segment responsible for the Tdi phenotype.
- To identify the genetic elements within this segment crucial for temperature-dependent instability.
Main Methods:
- Nucleotide sequencing of the Rts1 DNA b' segment.
- Expression analysis in Escherichia coli minicells.
- Construction and analysis of deletion derivatives.
- Site-directed mutagenesis.
Main Results:
- A 786-base-pair DNA segment responsible for Tdi was sequenced.
- An open reading frame (ORF) encoding a 16,000 molecular weight protein was identified within this segment.
- The chi sequence and a specific ORF (ORF I) were found to be essential for Tdi activity.
- Tdi was observed to be RecA-dependent but RecB-independent.
Conclusions:
- The Rts1 Tdi phenotype is mediated by a specific DNA segment containing a chi sequence, an ORF (ORF I), and a tail region.
- ORF I is critical for the expression of Tdi, suggesting a protein product is involved in the instability mechanism.