ArticleNucleic acids research2015
Highly efficient CRISPR/Cas9-mediated TAR cloning of genes and chromosomal loci from complex genomes in yeast.
Article in Nucleic acids research, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 69 papers.
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Who cites it
69 citing papers in PubMed.
- Mobile genetic elements drive a plasmid fusion and deletion lifecycle shaping evolution and antimicrobial resistance.Nature communications · 2026Article
- Yeast nuclei-mediated precise delivery of synthetic megabase-scale human DNA into mammalian embryos.Nature protocols · 2026Review
- Multi-Targeting Non-Specific Genome Engineering in Bacteria.Advanced science (Weinheim, Baden-Wurttemberg, Germany) · 2026Article
- iCASRED, a scarless DNA editing tool inSynthetic and systems biotechnology · 2025Article
- Enhancement of spinosad production in Saccharopolyspora spinosa by overexpression of the complete 74-kb spinosyn gene cluster.Microbial cell factories · 2025Article
- Current Approaches for Genetic Manipulation ofBiotech (Basel (Switzerland)) · 2025Review
- An Improved Transformation-Associated Recombination Cloning Approach for Direct Capturing of Natural Product Biosynthetic Gene Clusters.Microbial biotechnology · 2024Article
- Leveraging the power of long reads for targeted sequencing.Genome research · 2024Review
- A CRISPR-Cas9-Mediated Large-Fragment Assembly Method for Cloning Genomes and Biosynthetic Gene Cluster.Microorganisms · 2024Article
- A synthetic biology approach to assemble and reboot clinically relevantMicrobiology spectrum · 2024Article
- A simple method for rapid cloning of complete herpesvirus genomes.Cell reports methods · 2024Article
- Review
- Combining Fusion of Cells with CRISPR-Cas9 Editing for the Cloning of Large DNA Fragments or Complete Bacterial Genomes in Yeast.ACS synthetic biology · 2023Article
- Beyond assembly: the increasing flexibility of single-molecule sequencing technology.Nature reviews. Genetics · 2023Review
- High-titer production of staurosporine by heterologous expression and process optimization.Applied microbiology and biotechnology · 2023Article
- YLC-assembly: large DNA assembly via yeast life cycle.Nucleic acids research · 2023Article
- Applications of Programmable Endonucleases in Sequence- and Ligation-Independent Seamless DNA Assembly.Biomolecules · 2023Review
- High-throughput retrieval of target sequences from complex clone libraries using CRISPRi.Nature biotechnology · 2023Article
- Cross-species microbial genome transfer: a Review.Frontiers in bioengineering and biotechnology · 2023Review
- Chemical structure, properties and potential applications of surfactin, as well as advanced strategies for improving its microbial production.AIMS microbiology · 2023Review
9 more citing papers are in PubMed but not listed here.
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Authors and funding
3 authors.
Funding
Abstract
Transformation-associated recombination (TAR) protocol allowing the selective isolation of full-length genes complete with their distal enhancer regions and entire genomic loci with sizes up to 250 kb from complex genomes in yeast S. cerevisiae has been developed more than a decade ago. However, its wide spread usage has been impeded by a low efficiency (0.5-2%) of chromosomal region capture during yeast transformants which in turn requires a time-consuming screen of hundreds of colonies. Here, we demonstrate that pre-treatment of genomic DNA with CRISPR-Cas9 nucleases to generate double-strand breaks near the targeted genomic region results in a dramatic increase in the fraction of gene-positive colonies (up to 32%). As only a dozen or less yeast transformants need to be screened to obtain a clone with the desired chromosomal region, extensive experience with yeast is no longer required. A TAR-CRISPR protocol may help to create a bank of human genes, each represented by a genomic copy containing its native regulatory elements, that would lead to a significant advance in functional, structural and comparative genomics, in diagnostics, gene replacement, generation of animal models for human diseases and has a potential for gene therapy.
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