Linked Life Data

20967442

Source:http://linkedlifedata.com/resource/pubmed/id/20967442

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2011-1-4
pubmed:abstractText
Establishment of molecular diagnostics offering quantitative technology is directly associated with real-time polymerase chain reaction (PCR). This rapid, accurate and sensitive method requires careful execution, including reliable calibration standards. The storage of such standards is crucial to prevent nucleic acid decay and to ensure stable results using real-time PCR. In this study, a broad investigation of possible causes of DNA degradation during storage was performed, including GC-content of the fragments, long-term storage, rapid freeze-and-thaw experiments, genomic DNA and short DNA fragments of different species, the influence of shear stress and the effect of nuclease remaining after DNA isolation. Several known chemical DNA degradation mechanisms have been matched with the experimental data through a process of elimination. Protocols for practical application, as well as a theoretical model describing the underlying mechanisms of deviation of real-time PCR results due to decay of standard DNA, have been developed. Primary amines in the buffer composition, which enhance depurination of the DNA helix, and shear stress due to ice crystal formation, could be identified as major sources of interaction. This results in degradation of the standard DNA, as well as in the probability of occurrence of mismatches affecting real-time PCR performance.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
Jan
pubmed:issn
1432-0614
pubmed:author
pubmed:issnType
Electronic
pubmed:volume
89
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
407-17
pubmed:meshHeading
pubmed:year
2011
pubmed:articleTitle
Mechanisms of degradation of DNA standards for calibration function during storage.
pubmed:affiliation
Christian Doppler Laboratory for Molecular Food Analytics, Veterinärplatz 1, 1210 Vienna, Austria. peter.rossmanith@vetmeduni.ac.at
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't