ArticleBMC molecular biology2010
Single-stranded heteroduplex intermediates in lambda Red homologous recombination.
Article in BMC molecular biology, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 77 papers.
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77 citing papers in PubMed, 122 citations in OpenAlex.
- Development of a new recombineering system forSynthetic and systems biotechnology · 2027Article
- ETfinder: harnessing conserved C-terminal tails of single-stranded DNA-binding proteins for mining and engineering RecET systems in non-model microbial chassis.Nucleic acids research · 2026Article
- Enhancement of single-stranded template annealing activity by Rad52 during repair of CRISPR-induced dsDNA breaks.Nucleic acids research · 2026Article
- Development of an efficient recombineering system for Aeromonas.BMC microbiology · 2026Article
- RECKLEEN is a lambda Red/CRISPR-Cas9 based single plasmid platform for enhanced genome editing in Klebsiella pneumoniae.Communications biology · 2025Article
- iCASRED, a scarless DNA editing tool inSynthetic and systems biotechnology · 2025Article
- Synthetic and Functional Engineering of Bacteriophages: Approaches for Tailored Bactericidal, Diagnostic, and Delivery Platforms.Molecules (Basel, Switzerland) · 2025Review
- A diverse single-stranded DNA-annealing protein library enables efficient genome editing across bacterial phyla.Proceedings of the National Academy of Sciences of the United States of America · 2025Article
- Synthetic Biology in Natural Product Biosynthesis.Chemical reviews · 2025Review
- ReaL-MGE is a tool for enhanced multiplex genome engineering and application to malonyl-CoA anabolism.Nature communications · 2024Article
- Proof of ssDNA degraded from dsDNA for ET recombination.Biochemistry and biophysics reports · 2024Article
- ORBIT for E. coli: kilobase-scale oligonucleotide recombineering at high throughput and high efficiency.Nucleic acids research · 2024Article
- Half a century after their discovery: Structural insights into exonuclease and annealase proteins catalyzing recombineering.Engineering microbiology · 2024Review
- The emerging role of recombineering in microbiology.Engineering microbiology · 2023Review
- Single-Stranded DNA-Binding Proteins Mediate DSB Repair and Effectively Improve CRISPR/Cas9 Genome Editing inMicroorganisms · 2023Article
- Recombineering: Genetic Engineering in Escherichia coli Using Homologous Recombination.Current protocols · 2023Article
- RedβNature communications · 2022Article
- Robust counterselection and advanced λRed recombineering enable markerless chromosomal integration of large heterologous constructs.Nucleic acids research · 2022Article
- Improved dsDNA recombineering enables versatile multiplex genome engineering of kilobase-scale sequences in diverse bacteria.Nucleic acids research · 2022Article
- Generating Single Nucleotide Point Mutations in E. coli with the No-SCAR System.Methods in molecular biology (Clifton, N.J.) · 2022Article
17 more citing papers are in PubMed but not listed here.
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Authors and funding
6 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundThe Red proteins of lambda phage mediate probably the simplest and most efficient homologous recombination reactions yet described. However the mechanism of dsDNA recombination remains undefined.
resultsHere we show that the Red proteins can act via full length single stranded intermediates to establish single stranded heteroduplexes at the replication fork. We created asymmetrically digestible dsDNA substrates by exploiting the fact that Redalpha exonuclease activity requires a 5' phosphorylated end, or is blocked by phosphothioates. Using these substrates, we found that the most efficient configuration for dsDNA recombination occurred when the strand that can prime Okazaki-like synthesis contained both homology regions on the same ssDNA molecule. Furthermore, we show that Red recombination requires replication of the target molecule.
conclusionsHence we propose a new model for dsDNA recombination, termed 'beta' recombination, based on the formation of ssDNA heteroduplexes at the replication fork. Implications of the model were tested using (i) an in situ assay for recombination, which showed that recombination generated mixed wild type and recombinant colonies; and (ii) the predicted asymmetries of the homology arms, which showed that recombination is more sensitive to non-homologies attached to 5' than 3' ends. Whereas beta recombination can generate deletions in target BACs of at least 50 kb at about the same efficiency as small deletions, the converse event of insertion is very sensitive to increasing size. Insertions up to 3 kb are most efficiently achieved using beta recombination, however at greater sizes, an alternative Red-mediated mechanism(s) appears to be equally efficient. These findings define a new intermediate in homologous recombination, which also has practical implications for recombineering with the Red proteins.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.