ArticleNucleic acids research2012
A versatile element for gene addition in bacterial chromosomes.
Article in Nucleic acids research, 2012. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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13 citing papers in PubMed.
- Tunable low-rate genomic recombination with Cre-Applied and environmental microbiology · 2026Article
- Transposition with Tn3-family elements occurs through interaction with the host β-sliding clamp processivity factor.Nucleic acids research · 2024Article
- Novel mechanisms of diversity generation in Acinetobacter baumannii resistance islands driven by Tn7-like elements.Nucleic acids research · 2024Article
- Widespread CRISPR-derived RNA regulatory elements in CRISPR-Cas systems.Nucleic acids research · 2023Article
- Widespread CRISPR repeat-like RNA regulatory elements in CRISPR-Cas systems.bioRxiv : the preprint server for biology · 2023Article
- Discovery and characterization of novel type I-D CRISPR-guided transposons identified among diverse Tn7-like elements in cyanobacteria.Nucleic acids research · 2023Article
- inPOSE: A Flexible Toolbox for Chromosomal Cloning and Amplification of Bacterial Transgenes.Microorganisms · 2022Article
- Review
- Rpn (YhgA-Like) Proteins of Escherichia coli K-12 and Their Contribution to RecA-Independent Horizontal Transfer.Journal of bacteriology · 2017Article
- Long-Term Stable and Tightly Controlled Expression of Recombinant Proteins in Antibiotics-Free Conditions.PloS one · 2016Article
- Article
- Analysis of a taurine-dependent promoter in Sinorhizobium meliloti that offers tight modulation of gene expression.BMC microbiology · 2014Article
- Article
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2 authors.
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Abstract
The increasing interest in genetic manipulation of bacterial host metabolic pathways for protein or small molecule production has led to a need to add new genes to a chromosome quickly and easily without leaving behind a selectable marker. The present report describes a vector and four-day procedure that enable site-specific chromosomal insertion of cloned genes in a context insulated from external transcription, usable once in a construction series. The use of rhamnose-inducible transcription from rhaBp allows regulation of the inserted genes independently of the commonly used IPTG and arabinose strategies. Using lacZ as a reporter, we first show that expression from the rhamnose promoter is tightly regulatable, exhibiting very low leakage of background expression compared with background, and moderate rhamnose-induced expression compared with IPTG-induced expression from lacp. Second, the expression of a DNA methyltransferase was used to show that rhamnose regulation yielded on-off expression of this enzyme, such that a resident high-copy plasmid was either fully sensitive or fully resistant to isoschizomer restriction enzyme cleavage. In both cases, growth medium manipulation allows intermediate levels of expression. The vehicle can also be adapted as an ORF-cloning vector.
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