| Separation and recovery of nucleic acids with improved biological activity by acid-degradable polyacrylamide gel electrophoresis. | |
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MedLine Citation:
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PMID: 20419706 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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One of the fundamental challenges in studying biomacromolecules (e.g. nucleic acids and proteins) and their complexes in a biological system is isolating them in their structurally and functionally intact forms. Electrophoresis offers convenient and efficient separation and analysis of biomacromolecules but recovery of separated biomacromolecules is a significant challenge. In this study, DNAs of various sizes were separated by electrophoresis in an acid-degradable polyacrylamide gel. Almost 100% of the nucleic acids were recovered after the identified gel bands were hydrolyzed under a mildly acidic condition and purified using anion exchange resin. Further concentration by centrifugal filtration and a second purification using ion exchange column chromatography yielded 44-84% of DNA. The second conventional (non-degradable) gel electrophoresis confirmed that the nucleic acids recovered from acid-degradable gel bands preserved their electrophoretic properties through acidic gel hydrolysis, purification, and concentration processes. The plasmid DNA recovered from acid-degradable gel transfected cells significantly more efficiently than the starting plasmid DNA (i.e. improved biological activity via acid-degradable PAGE). Separation of other types of nucleic acids such as small interfering RNA using this convenient and efficient technique was also demonstrated. |
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Authors:
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Yoon Kyung Kim; Young Jik Kwon |
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Publication Detail:
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Type: Journal Article |
Journal Detail:
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Title: Electrophoresis Volume: 31 ISSN: 1522-2683 ISO Abbreviation: Electrophoresis Publication Date: 2010 May |
Date Detail:
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Created Date: 2010-05-25 Completed Date: 2010-08-30 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 8204476 Medline TA: Electrophoresis Country: Germany |
Other Details:
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Languages: eng Pagination: 1656-61 Citation Subset: IM |
Affiliation:
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Department of Biomedical Engineering, University of California, Irvine, CA, USA. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Animals DNA / isolation & purification Electrophoresis, Polyacrylamide Gel / methods* Green Fluorescent Proteins Hydrogen-Ion Concentration Mice Microscopy, Fluorescence NIH 3T3 Cells Nucleic Acids / isolation & purification* Plasmids / isolation & purification RNA, Small Interfering / isolation & purification Transfection |
| Chemical | |
Reg. No./Substance:
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0/Nucleic Acids; 0/RNA, Small Interfering; 147336-22-9/Green Fluorescent Proteins; 9007-49-2/DNA |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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