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Article: Efficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli

TitleEfficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia coli
Authors
Issue Date2005
PublisherOxford University Press. The Journal's web site is located at http://nar.oxfordjournals.org/
Citation
Nucleic Acids Research, 2005, v. 33 n. 6, p. 1-9 How to Cite?
Abstract
λ-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.
Persistent Identifierhttp://hdl.handle.net/10722/48990
ISSN
2013 Impact Factor: 8.808
PubMed Central ID
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorWong, QNYen_HK
dc.contributor.authorNg, VCWen_HK
dc.contributor.authorLin, MCMen_HK
dc.contributor.authorKung, HFen_HK
dc.contributor.authorChan, Den_HK
dc.contributor.authorHuang, JDen_HK
dc.date.accessioned2008-06-12T06:31:31Z-
dc.date.available2008-06-12T06:31:31Z-
dc.date.issued2005en_HK
dc.identifier.citationNucleic Acids Research, 2005, v. 33 n. 6, p. 1-9en_HK
dc.identifier.issn0305-1048en_HK
dc.identifier.urihttp://hdl.handle.net/10722/48990-
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.en_HK
dc.format.extent388 bytes-
dc.format.mimetypetext/html-
dc.languageengen_HK
dc.publisherOxford University Press. The Journal's web site is located at http://nar.oxfordjournals.org/en_HK
dc.relation.ispartofNucleic Acids Researchen_HK
dc.rightsCreative Commons: Attribution 3.0 Hong Kong License-
dc.subject.meshDNA, Recombinant - geneticsen_HK
dc.subject.meshEscherichia coli - enzymology - genetics - growth & developmenten_HK
dc.subject.meshGenetic Engineering - methodsen_HK
dc.subject.meshRecombination, Geneticen_HK
dc.subject.meshThymidylate Synthase - geneticsen_HK
dc.titleEfficient and seamless DNA recombineering using a thymidylate synthase A selection system in Escherichia colien_HK
dc.typeArticleen_HK
dc.identifier.openurlhttp://library.hku.hk:4550/resserv?sid=HKU:IR&issn=0305-1048&volume=33&issue=6&spage=e59&epage=&date=2005&atitle=Efficient+and+seamless+DNA+recombineering+using+a+thymidylate+synthase+A+selection+system+in+Escherichia+colien_HK
dc.identifier.emailLin, MCM:mcllin@hkucc.hku.hken_HK
dc.identifier.emailChan, D:chand@hkucc.hku.hken_HK
dc.identifier.emailHuang, JD:jdhuang@hkucc.hku.hken_HK
dc.identifier.authorityLin, MCM=rp00746en_HK
dc.identifier.authorityChan, D=rp00540en_HK
dc.identifier.authorityHuang, JD=rp00451en_HK
dc.description.naturepublished_or_final_versionen_HK
dc.identifier.doi10.1093/nar/gni059en_HK
dc.identifier.pmid15800210en_HK
dc.identifier.pmcidPMC1072810en_HK
dc.identifier.scopuseid_2-s2.0-28644449037en_HK
dc.identifier.hkuros98288-
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-28644449037&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume33en_HK
dc.identifier.issue6en_HK
dc.identifier.spage1en_HK
dc.identifier.epage9en_HK
dc.identifier.eissn1362-4962-
dc.identifier.isiWOS:000228398400007-
dc.publisher.placeUnited Kingdomen_HK
dc.identifier.scopusauthoridWong, QNY=9742981900en_HK
dc.identifier.scopusauthoridNg, VCW=8215749500en_HK
dc.identifier.scopusauthoridLin, MCM=7404816359en_HK
dc.identifier.scopusauthoridKung, HF=7402514190en_HK
dc.identifier.scopusauthoridChan, D=7402216545en_HK
dc.identifier.scopusauthoridHuang, JD=8108660600en_HK
dc.identifier.citeulike161569-

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