ArticleNucleic acids research2018
ExoCET: exonuclease in vitro assembly combined with RecET recombination for highly efficient direct DNA cloning from complex genomes.
Article in Nucleic acids research, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. An erratum has been issued. Cited by 75 papers, 1 of them a synthesis that pooled it.
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Who cites it
75 citing papers in PubMed, 1 synthesis or guideline pooled it, 148 citations in OpenAlex.
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- Development of a bacterial gene transcription activating strategy based on transcriptional activator positive feedback.Journal of advanced research · 2024Article
- Seamless site-directed mutagenesis in complex cloned DNA sequences using the RedEx method.Nature protocols · 2024Review
- CRISETR: an efficient technology for multiplexed refactoring of biosynthetic gene clusters.Nucleic acids research · 2024Article
15 more citing papers are in PubMed but not listed here.
Corrections and comments
- Erratum issued
Authors and funding
11 authors at 5 institutions in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The exponentially increasing volumes of DNA sequence data highlight the need for new DNA cloning methods to explore the new information. Here, we describe 'ExoCET' (Exonuclease Combined with RecET recombination) to directly clone any chosen region from bacterial and mammalian genomes with nucleotide precision into operational plasmids. ExoCET combines in vitro exonuclease and annealing with the remarkable capacity of full length RecET homologous recombination (HR) to retrieve specified regions from genomic DNA preparations. Using T4 polymerase (T4pol) as the in vitro exonuclease for ExoCET, we directly cloned large regions (>50 kb) from bacterial and mammalian genomes, including DNA isolated from blood. Employing RecET HR or Cas9 cleavage in vitro, the directly cloned region can be chosen with nucleotide precision to position, for example, a gene into an expression vector without the need for further subcloning. In addition to its utility for bioprospecting in bacterial genomes, ExoCET presents straightforward access to mammalian genomes for various applications such as region-specific DNA sequencing that retains haplotype phasing, the rapid construction of optimal, haplotypic, isogenic targeting constructs or a new way to genotype that presents advantages over Southern blotting or polymerase chain reaction. The direct cloning capacities of ExoCET present new freedoms in recombinant DNA technology.
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