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Article: One-step DNA fragment assembly and circularization for gene cloning.

TitleOne-step DNA fragment assembly and circularization for gene cloning.
Authors
Issue Date2010
Citation
Current Issues In Molecular Biology, 2010, v. 12 n. 1, p. 11-16 How to Cite?
AbstractThis article describes a one-step procedure based on Taq polymerase for the precise assembly of DNA fragments into circular constructs as long as 6 kb. The only prior step needed was the amplification of the gene to be cloned and the linear vector backbone, and the whole process up to assembly and circularization lasted only 2 days, compared with the conventional method's 2 weeks. Furthermore, the final DNA construct was used to transform Escherichia coli directly without any further treatment. By circumventing the need for DNA ligase, our "Quick Assemble" method offers an improvement over the combination of long PCR and overlap extension PCR, and is expected to facilitate various kinds of complex genetic engineering projects that require precise in-frame assembly of multiple fragments, such as multiple site-directed mutagenesis and whole-DNA library gene shuffling, as well as the construction of new plasmids with any promoter, resistance gene marker, restriction site, or any DNA tag.
Persistent Identifierhttp://hdl.handle.net/10722/154618
ISSN
2015 SCImago Journal Rankings: 1.461
ISI Accession Number ID

 

DC FieldValueLanguage
dc.contributor.authorZuo, Pen_US
dc.contributor.authorRabie, BMen_US
dc.date.accessioned2012-08-08T08:26:31Z-
dc.date.available2012-08-08T08:26:31Z-
dc.date.issued2010en_US
dc.identifier.citationCurrent Issues In Molecular Biology, 2010, v. 12 n. 1, p. 11-16en_US
dc.identifier.issn1467-3045en_US
dc.identifier.urihttp://hdl.handle.net/10722/154618-
dc.description.abstractThis article describes a one-step procedure based on Taq polymerase for the precise assembly of DNA fragments into circular constructs as long as 6 kb. The only prior step needed was the amplification of the gene to be cloned and the linear vector backbone, and the whole process up to assembly and circularization lasted only 2 days, compared with the conventional method's 2 weeks. Furthermore, the final DNA construct was used to transform Escherichia coli directly without any further treatment. By circumventing the need for DNA ligase, our "Quick Assemble" method offers an improvement over the combination of long PCR and overlap extension PCR, and is expected to facilitate various kinds of complex genetic engineering projects that require precise in-frame assembly of multiple fragments, such as multiple site-directed mutagenesis and whole-DNA library gene shuffling, as well as the construction of new plasmids with any promoter, resistance gene marker, restriction site, or any DNA tag.en_US
dc.languageengen_US
dc.relation.ispartofCurrent issues in molecular biologyen_US
dc.subject.meshAnimalsen_US
dc.subject.meshBlotting, Westernen_US
dc.subject.meshCell Line, Tumoren_US
dc.subject.meshCloning, Molecular - Methodsen_US
dc.subject.meshDna - Chemistry - Geneticsen_US
dc.subject.meshDna, Complementary - Geneticsen_US
dc.subject.meshEscherichia Coli - Geneticsen_US
dc.subject.meshMiceen_US
dc.subject.meshModels, Geneticen_US
dc.subject.meshTransformation, Genetic - Geneticsen_US
dc.subject.meshAlpha-Synuclein - Genetics - Metabolismen_US
dc.titleOne-step DNA fragment assembly and circularization for gene cloning.en_US
dc.typeArticleen_US
dc.identifier.emailZuo, P:peijunzuo@lycos.comen_US
dc.identifier.emailRabie, BM:rabie@hku.hken_US
dc.identifier.authorityZuo, P=rp00050en_US
dc.identifier.authorityRabie, BM=rp00029en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.pmid19494420-
dc.identifier.scopuseid_2-s2.0-77954129380en_US
dc.identifier.hkuros156041-
dc.identifier.volume12en_US
dc.identifier.issue1en_US
dc.identifier.spage11en_US
dc.identifier.epage16en_US
dc.identifier.isiWOS:000269076300002-
dc.identifier.scopusauthoridZuo, P=36490907500en_US
dc.identifier.scopusauthoridRabie, BM=7007172734en_US

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