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Article: Efficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli
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TitleEfficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli
 
AuthorsWong, QNY1
Ng, VCW1
Lin, MCM1
Kung, HF1
Chan, D1
Huang, JD1
 
Issue Date2005
 
PublisherOxford University Press. The Journal's web site is located at http://nar.oxfordjournals.org/
 
CitationNucleic Acids Research, 2005, v. 33 n. 6, p. 1-9 [How to Cite?]
DOI: http://dx.doi.org/10.1093/nar/gni059
 
Abstractλ-Red system-based recombinogenic engineering is a powerful new method to engineer DNA without the need for restriction enzymes or ligases. Here, we report the use of a single selectable marker to enhance the usefulness of this approach. The strategy is to utilize the thymidylate synthase A (thyA) gene, which encodes an enzyme involved in the synthesis of thymidine 5′-triphosphate, for both positive and negative selection. With this approach, we successfully created point mutations in plasmid and bacterial artificial chromosome (BAC) DNA containing the mouse Col10a1 gene. The results showed that the thyA selection system is highly efficient and accurate, giving an average of >90% selection efficiency. This selection system produces DNA that is free from permanent integration of unwanted sequences, thus allowing unlimited rounds of modifications if required. © The Author 2005. Published by Oxford University Press. All rights reserved.
 
ISSN0305-1048
2013 Impact Factor: 8.808
 
DOIhttp://dx.doi.org/10.1093/nar/gni059
 
PubMed Central IDPMC1072810
 
ISI Accession Number IDWOS:000228398400007
 
ReferencesReferences in Scopus
 
DC FieldValue
dc.contributor.authorWong, QNY
 
dc.contributor.authorNg, VCW
 
dc.contributor.authorLin, MCM
 
dc.contributor.authorKung, HF
 
dc.contributor.authorChan, D
 
dc.contributor.authorHuang, JD
 
dc.date.accessioned2008-06-12T06:31:31Z
 
dc.date.available2008-06-12T06:31:31Z
 
dc.date.issued2005
 
dc.description.abstractλ-Red system-based recombinogenic engineering is a powerful new method to engineer DNA without the need for restriction enzymes or ligases. Here, we report the use of a single selectable marker to enhance the usefulness of this approach. The strategy is to utilize the thymidylate synthase A (thyA) gene, which encodes an enzyme involved in the synthesis of thymidine 5′-triphosphate, for both positive and negative selection. With this approach, we successfully created point mutations in plasmid and bacterial artificial chromosome (BAC) DNA containing the mouse Col10a1 gene. The results showed that the thyA selection system is highly efficient and accurate, giving an average of >90% selection efficiency. This selection system produces DNA that is free from permanent integration of unwanted sequences, thus allowing unlimited rounds of modifications if required. © The Author 2005. Published by Oxford University Press. All rights reserved.
 
dc.description.naturepublished_or_final_version
 
dc.format.extent388 bytes
 
dc.format.mimetypetext/html
 
dc.identifier.citationNucleic Acids Research, 2005, v. 33 n. 6, p. 1-9 [How to Cite?]
DOI: http://dx.doi.org/10.1093/nar/gni059
 
dc.identifier.citeulike161569
 
dc.identifier.doihttp://dx.doi.org/10.1093/nar/gni059
 
dc.identifier.eissn1362-4962
 
dc.identifier.epage9
 
dc.identifier.hkuros98288
 
dc.identifier.isiWOS:000228398400007
 
dc.identifier.issn0305-1048
2013 Impact Factor: 8.808
 
dc.identifier.issue6
 
dc.identifier.openurl
 
dc.identifier.pmcidPMC1072810
 
dc.identifier.pmid15800210
 
dc.identifier.scopuseid_2-s2.0-28644449037
 
dc.identifier.spage1
 
dc.identifier.urihttp://hdl.handle.net/10722/48990
 
dc.identifier.volume33
 
dc.languageeng
 
dc.publisherOxford University Press. The Journal's web site is located at http://nar.oxfordjournals.org/
 
dc.publisher.placeUnited Kingdom
 
dc.relation.ispartofNucleic Acids Research
 
dc.relation.referencesReferences in Scopus
 
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License
 
dc.subject.meshDNA, Recombinant - genetics
 
dc.subject.meshEscherichia coli - enzymology - genetics - growth & development
 
dc.subject.meshGenetic Engineering - methods
 
dc.subject.meshRecombination, Genetic
 
dc.subject.meshThymidylate Synthase - genetics
 
dc.titleEfficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli
 
dc.typeArticle
 
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Author Affiliations
  1. The University of Hong Kong