ArticleNucleic acids research2008
Recombineering with tolC as a selectable/counter-selectable marker: remodeling the rRNA operons of Escherichia coli.
Article in Nucleic acids research, 2008. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 47 papers.
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Who cites it
47 citing papers in PubMed, 74 citations in OpenAlex.
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- Recombineering: Genetic Engineering in Escherichia coli Using Homologous Recombination.Current protocols · 2023Article
- Efficient Bacterial Genome Engineering throughout the Central Dogma Using the Dual-Selection MarkerACS synthetic biology · 2022Article
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- Bacterial Genetic Engineering by Means of Recombineering for Reverse Genetics.Frontiers in microbiology · 2020Review
- An Engineered Cas-Transposon System for Programmable and Site-Directed DNA Transpositions.The CRISPR journal · 2019Article
- In vivo continuous evolution of metabolic pathways for chemical production.Microbial cell factories · 2019Review
- FlexiBAC: a versatile, open-source baculovirus vector system for protein expression, secretion, and proteolytic processing.BMC biotechnology · 2019Article
- Characterization of Inducible ccdB Gene as a Counterselectable Marker in Escherichia coli Recombineering.Current microbiology · 2017Article
- Transcriptomic analysis displays the effect of (-)-roemerine on the motility and nutrient uptake in Escherichia coli.Current genetics · 2017Article
- Oligo- and dsDNA-mediated genome editing using a tetA dual selection system in Escherichia coli.PloS one · 2017Article
- λ Recombination and Recombineering.EcoSal Plus · 2016Review
- Systems Metabolic Engineering of Escherichia coli.EcoSal Plus · 2016Review
- Engineering an allosteric transcription factor to respond to new ligands.Nature methods · 2016Article
Corrections and comments
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Authors and funding
1 author at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
This work describes the novel use of tolC as a selectable/counter-selectable marker for the facile modification of DNA in Escherichia coli. Expression of TolC (an outer membrane protein) confers relative resistance to toxic small molecules, while its absence renders the cell tolerant to colicin E1. These features, coupled with the lambdaredgam recombination system, allow for selection of tolC insertions/deletions anywhere on the E. coli chromosome or on plasmid DNA. This methodology obviates the need for minimal growth media, specialized wash protocols and the lengthy incubation times required by other published recombineering methods. As a rigorous test of the TolC selection system, six out of seven 23S rRNA genes were consecutively and seamlessly removed from the E. coli chromosome without affecting expression of neighboring genes within the complex rrn operons. The resulting plasmid-free strain retains one 23S rRNA gene (rrlC) in its natural location on the chromosome and is the first mutant of its kind. These new rRNA mutants will be useful in the study of rRNA gene regulation and ribosome function. Given its high efficiency, low background and facility in rich media, tolC selection is a broadly applicable method for the modification of DNA by recombineering.
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Registered trials
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