Linked Life Data

9826780

Source:http://linkedlifedata.com/resource/pubmed/id/9826780

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
23
pubmed:dateCreated
1999-2-3
pubmed:abstractText
Here we analyze the effect of DNA folding on the performance of short primers and describe a simple technique for assessing hitherto uncertain values of thermodynamic parameters that determine the folding of single-stranded DNA into secondary structure. An 8mer with two degenerate positions is extended simultaneously at several complementary sites on a known template (M13mp18) using one, two or three (but never all four) of the possible dNTPs. The length of the extension is site specific because it is limited by the first occurrence in the downstream template sequence of a base whose complementary dNTP is not present. The relative priming efficiencies of different sites are then ranked by comparing their band brightnesses on a gel. The priming efficiency of a short primer (unlike conventional long primers) depends dramatically on the secondary structure of the template at and around the priming site. We calculated the secondary structure and its effect on priming using a simple model with relatively few parameters which were then optimized to achieve the best match between the predictions and the actual rankings of the sites in terms of priming efficiency. This work introduces an efficient and conceptually novel approach that in the future can make use of more data to optimize a larger set of DNA folding parameters in a more refined model. The model we used, however crude it may be, significantly improved the prediction of priming efficiencies of 8mer primers and appreciably raised the success rate of our DNA sequencing technique (from 67 to 91% with a significance of P < 7 x 10(-5)), which uses such primers.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Dec
pubmed:issn
0305-1048
pubmed:author
pubmed:issnType
Print
pubmed:day
1
pubmed:volume
26
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
5525-32
pubmed:dateRevised
2008-11-20
pubmed:meshHeading
pubmed:year
1998
pubmed:articleTitle
Interdependence between DNA template secondary structure and priming efficiencies of short primers.
pubmed:affiliation
Department of Structural Biology, Weizmann Institute of Science, Rehovot 76100, Israel. levy@anl.gov
pubmed:publicationType
Journal Article, Research Support, U.S. Gov't, Non-P.H.S.