ArticleGenome research2002
Linker-mediated recombinational subcloning of large DNA fragments using yeast.
Article in Genome research, 2002. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 28 papers.
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Who cites it
28 citing papers in PubMed.
- Wzx flippases exhibiting complex O-unit preferences require a new model for Wzx-substrate interactions.MicrobiologyOpen · 2019Article
- Improving acetyl-CoA biosynthesis inBiotechnology for biofuels · 2017Article
- Scaffolded Antigens in Yeast Cell Particle Vaccines Provide Protection against Systemic Polyoma Virus Infection.Journal of immunology research · 2016Article
- Simultaneous knockdown of six non-family genes using a single synthetic RNAi fragment in Arabidopsis thaliana.Plant methods · 2016Article
- A versatile, efficient strategy for assembly of multi-fragment expression vectors in Saccharomyces cerevisiae using 60 bp synthetic recombination sequences.Microbial cell factories · 2013Article
- Ultra-low background DNA cloning system.PloS one · 2013Article
- Direct cloning of isogenic murine DNA in yeast and relevance of isogenicity for targeting in embryonic stem cells.PloS one · 2013Article
- New vector tools with a hygromycin resistance marker for use with opportunistic pathogens.Molecular biotechnology · 2011Article
- Cloning of the repertoire of individual Plasmodium falciparum var genes using transformation associated recombination (TAR).PloS one · 2011Article
- Introduction of large DNA inserts into the barley pathogenic fungus, Ustilago hordei, via recombined binary BAC vectors and Agrobacterium-mediated transformation.Current genetics · 2011Article
- Oligomeric size of the m2 muscarinic receptor in live cells as determined by quantitative fluorescence resonance energy transfer.The Journal of biological chemistry · 2010Article
- Phosphorylation of the conserved transcription factor ATF-7 by PMK-1 p38 MAPK regulates innate immunity in Caenorhabditis elegans.PLoS genetics · 2010Article
- Synthesis of DNA fragments in yeast by one-step assembly of overlapping oligonucleotides.Nucleic acids research · 2009Article
- New yeast recombineering tools for bacteria.Plasmid · 2009Article
- DNA assembler, an in vivo genetic method for rapid construction of biochemical pathways.Nucleic acids research · 2009Article
- One-step assembly in yeast of 25 overlapping DNA fragments to form a complete synthetic Mycoplasma genitalium genome.Proceedings of the National Academy of Sciences of the United States of America · 2008Article
- Efficient transfer of two large secondary metabolite pathway gene clusters into heterologous hosts by transposition.Nucleic acids research · 2008Article
- Yeast-based recombineering of DNA fragments into plant transformation vectors by one-step transformation.Plant cell reports · 2007Article
- Plasmodium falciparum: a novel method for analyzing haplotypes in mixed infections.Experimental parasitology · 2007Article
- In vivo construction of recombinant molecules within the Caenorhabditis elegans germ line using short regions of terminal homology.Nucleic acids research · 2007Article
Corrections and comments
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Authors and funding
3 authors.
Funding
Abstract
The homologous recombination pathway in yeast is an ideal tool for the sequence-specific assembly of plasmids. Complementary 80-nucleotide oligonucleotides that overlap a vector and a target fragment were found to serve as efficient recombination linkers for fragment subcloning. Using electroporation, single-stranded 80-mers were adequate for routine plasmid construction. A cycloheximide-based counterselection was introduced to increase the specificity of cloning by homologous recombination relative to nonspecific vector background. Reconstruction experiments suggest this counterselection increased cloning specificity by 100-fold. Cycloheximide counterselection was used in conjunction with 80-bp linkers to subclone targeted regions from bacterial artificial chromosomes. This technology may find broad application in the final stages of completing the Human Genome Sequencing Project and in applications of BAC clones to the functional analysis of complex genomes.
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Registered trials
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.