Linked Life Data

15585138

Source:http://linkedlifedata.com/resource/pubmed/id/15585138

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
11
pubmed:dateCreated
2004-12-8
pubmed:abstractText
The effect of urea, isopropanol, propan-1-ol, and butan-1-ol on PCR using three different DNA polymerases was investigated. In the presence of these agents, polymerases were active as expected up to a critical concentration where they became progressively inhibited. Critical concentrations of alcohols generally increased with thermoresistance of the polymerases and decreased with the hydrophobicity of the alcohols. These results indicate that an important aspect of the inhibition involved conformational loosening due to a decrease in the hydrophobic effect. A mutagenic effect occurred with Vent(r) (exo-) DNA polymerase in the presence of 7.0 to 8.0% v/v propan-1-ol, affording mutation frequencies of up to 9.8 x 10(-3) mutation/bp/PCR. Under these conditions the preferential replacement of Gs and Cs was observed, in opposition to standard error-prone PCR that favors replacement of As and Ts. Comparison of various PCR conditions indicates that propanol and MnCl2 have different modes of action, and that the decrease in fidelity promoted by propanol is due to a finely tuned partial destabilization of the polymerase. The PCR conditions developed in this study provide a useful alternative for targeting different sequence space for directed evolution experiments.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Nov
pubmed:issn
1044-5498
pubmed:author
pubmed:issnType
Print
pubmed:volume
23
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
789-95
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2004
pubmed:articleTitle
Alcohol-mediated error-prone PCR.
pubmed:affiliation
Groupe de Recherche en Energie et Information Biomoléculaires, Université du Québec à Trois-Rivières, Trois-Rivières, Québec, Canada.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't