Two-dimensional conformation-dependent electrophoresis (2D-CDE) to separate DNA fragments containing unmatched bulge from complex DNA samples

2.50
Hdl Handle:
http://hdl.handle.net/2336/3457
Title:
Two-dimensional conformation-dependent electrophoresis (2D-CDE) to separate DNA fragments containing unmatched bulge from complex DNA samples
Authors:
Gunnarsson, Gudmundur H; Thormar, Hans G; Gudmundsson, Bjarki; Akesson, Lina; Jonsson, Jon J
Citation:
Nucleic Acids Res. 2004, 32(2):e23
Issue Date:
2004
Abstract:
DNA fragments containing mispaired and modified bases, bulges, lesions and specific sequences have altered conformation. Methods for separating complex samples of DNA fragments based on conformation but independent of length have many applications, including (i) separation of mismatched or unmatched DNA fragments from those perfectly matched; (ii) simultaneous, diagnostic, mismatch scanning of multiple fragments; (iii) isolation of damaged DNA fragments from undamaged fragments; and (iv) estimation of reannealing efficiency of complex DNA samples. We developed a two-dimensional conformation-dependent electrophoresis (2D-CDE) method for separating DNA fragments based on length and conformation in the first dimension and only on length in the second dimension. Differences in migration velocity due to conformation were minimized during second dimension electrophoresis by introducing an intercalator. To test the method, we constructed 298 bp DNA fragments containing cytosine bulges ranging from 1 to 5 nt. Bulge-containing DNA fragments had reduced migration velocity in the first dimension due to altered conformation. After 2D-CDE, bulge-containing DNA fragments had migrated in front of an arc comprising heterogeneous fragments with regular conformation. This simple and robust method could be used in both analytical and preparative applications involving complex DNA samples.
Description:
To access publisher full text version of this article. Please click on the hyperlink in Additional Links field
Additional Links:
http://nar.oxfordjournals.org/cgi/content/full/32/2/e23

Full metadata record

DC FieldValue Language
dc.contributor.authorGunnarsson, Gudmundur H-
dc.contributor.authorThormar, Hans G-
dc.contributor.authorGudmundsson, Bjarki-
dc.contributor.authorAkesson, Lina-
dc.contributor.authorJonsson, Jon J-
dc.date.accessioned2006-07-14T12:53:12Z-
dc.date.available2006-07-14T12:53:12Z-
dc.date.issued2004-
dc.identifier.citationNucleic Acids Res. 2004, 32(2):e23en
dc.identifier.issn1362-4962-
dc.identifier.pmid14762200-
dc.identifier.doi10.1093/nar/gnh018-
dc.identifier.urihttp://hdl.handle.net/2336/3457-
dc.descriptionTo access publisher full text version of this article. Please click on the hyperlink in Additional Links fielden
dc.description.abstractDNA fragments containing mispaired and modified bases, bulges, lesions and specific sequences have altered conformation. Methods for separating complex samples of DNA fragments based on conformation but independent of length have many applications, including (i) separation of mismatched or unmatched DNA fragments from those perfectly matched; (ii) simultaneous, diagnostic, mismatch scanning of multiple fragments; (iii) isolation of damaged DNA fragments from undamaged fragments; and (iv) estimation of reannealing efficiency of complex DNA samples. We developed a two-dimensional conformation-dependent electrophoresis (2D-CDE) method for separating DNA fragments based on length and conformation in the first dimension and only on length in the second dimension. Differences in migration velocity due to conformation were minimized during second dimension electrophoresis by introducing an intercalator. To test the method, we constructed 298 bp DNA fragments containing cytosine bulges ranging from 1 to 5 nt. Bulge-containing DNA fragments had reduced migration velocity in the first dimension due to altered conformation. After 2D-CDE, bulge-containing DNA fragments had migrated in front of an arc comprising heterogeneous fragments with regular conformation. This simple and robust method could be used in both analytical and preparative applications involving complex DNA samples.en
dc.language.isoenen
dc.publisherOxford University Pressen
dc.relation.urlhttp://nar.oxfordjournals.org/cgi/content/full/32/2/e23en
dc.subjectCytosineen
dc.subjectDNAen
dc.subjectElectrophoresis, Gel, Two-Dimensionalen
dc.subjectEthidiumen
dc.subjectHeteroduplex Analysisen
dc.subjectNucleic Acid Conformationen
dc.subjectNucleic Acid Heteroduplexesen
dc.subjectReproducibility of Resultsen
dc.subjectResearch Support, Non-U.S. Gov'ten
dc.subjectSensitivity and Specificityen
dc.subjectTemperatureen
dc.titleTwo-dimensional conformation-dependent electrophoresis (2D-CDE) to separate DNA fragments containing unmatched bulge from complex DNA samplesen
dc.typeArticleen
dc.contributor.departmentDepartment of Biochemistry and Molecular Biology, Faculty of Medicine, University of Iceland, IS-101 Reykjavik, Iceland.is
dc.identifier.journalNucleic acids researchen
dc.format.digYES-

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