Linked Life Data

10339550

Source:http://linkedlifedata.com/resource/pubmed/id/10339550

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
11
pubmed:dateCreated
1999-6-24
pubmed:abstractText
We report a general method for screening, in solution, the impact of deviations from canonical Watson-Crick composition on the thermodynamic stability of nucleic acid duplexes. We demonstrate how fluorescence resonance energy transfer (FRET) can be used to detect directly free energy differences between an initially formed "reference" duplex (usually a Watson-Crick duplex) and a related "test" duplex containing a lesion/alteration of interest (e.g., a mismatch, a modified, a deleted, or a bulged base, etc.). In one application, one titrates into a solution containing a fluorescently labeled, FRET-active, reference duplex, an unlabeled, single-stranded nucleic acid (test strand), which may or may not compete successfully to form a new duplex. When a new duplex forms by strand displacement, it will not exhibit FRET. The resultant titration curve (normalized fluorescence intensity vs. logarithm of test strand concentration) yields a value for the difference in stability (free energy) between the newly formed, test strand-containing duplex and the initial reference duplex. The use of competitive equilibria in this assay allows the measurement of equilibrium association constants that far exceed the magnitudes accessible by conventional titrimetric techniques. Additionally, because of the sensitivity of fluorescence, the method requires several orders of magnitude less material than most other solution methods. We discuss the advantages of this method for detecting and characterizing any modification that alters duplex stability, including, but not limited to, mutagenic lesions. We underscore the wide range of accessible free energy values that can be defined by this method, the applicability of the method in probing for a myriad of nucleic acid variations, such as single nucleotide polymorphisms, and the potential of the method for high throughput screening.
pubmed:grant
pubmed:commentsCorrections
http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-1406315, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-2271593, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-2440861, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-4573844, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-8092707, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-8519772, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-8538456, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-9170325, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-9585546, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-9730823, http://linkedlifedata.com/resource/pubmed/commentcorrection/10339550-9760268
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
0027-8424
pubmed:author
pubmed:issnType
Print
pubmed:day
25
pubmed:volume
96
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
6113-8
pubmed:dateRevised
2009-11-18
pubmed:meshHeading
pubmed:year
1999
pubmed:articleTitle
A quantitative method for evaluating the stabilities of nucleic acids.
pubmed:affiliation
Department of Chemistry, Rutgers, The State University of New Jersey, 610 Taylor Road, Piscataway, NJ 08854, USA.
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, P.H.S.