Linked Life Data

10785287

Source:http://linkedlifedata.com/resource/pubmed/id/10785287

Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
2
pubmed:dateCreated
2000-6-30
pubmed:abstractText
Test conditions for the simultaneous detection of potato leafroll virus (PLRV) and potato virus Y (PVY) in dormant tubers and leaves by reverse transcription-polymerase chain reaction (RT-PCR) were optimized. Various factors optimized at the reverse transcription (RT) stage rather than at the amplification (PCR) stage affected the outcome. In the simplex RT-PCR a onefold dNTPs concentration (0.5 mM) was sufficient in yielding a PLRV or PVY band. In contrast, the duplex RT-PCR required a minimum twofold dNTPs concentration (1.0 mM) during RT to produce distinct bands in PCR. Similarly, various proportions of antisense primers of PLRV and PVY used during RT affected subsequent duplex RT-PCR. Optimal amplification of both viruses were obtained at a ratio of 0.90:0.49 microM of PLRV:PVY antisense primers. An interaction of dNTPs and RNA template concentration was observed. A higher concentration of RNA was required at onefold dNTPs concentration than at twofold dNTPs. Dilutions down to 1:300 of RNA template yielded distinct bands of both viruses at twofold dNTPs concentration. At optimized conditions of the duplex RT-PCR both viruses were reliably detected in composite samples at a ratio of one part infected sap mixed with 399 parts of sap from healthy tubers. Application of optimized conditions to singly- and doubly-infected tubers detected both viruses from naturally infected field-grown tubers. A nearly perfect correlation (r(2)=0.99) was observed between visible plant symptoms and the virus detection from leaves and tubers by the duplex RT-PCR.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
pubmed:status
MEDLINE
pubmed:month
May
pubmed:issn
0166-0934
pubmed:author
pubmed:issnType
Print
pubmed:volume
86
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
121-9
pubmed:dateRevised
2006-11-15
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
Duplex RT-PCR: reagent concentrations at reverse transcription stage affect the PCR performance.
pubmed:affiliation
Potato Research Centre, Agriculture and Agri-Food Canada, P.O. Box 20280, Fredericton, Canada. singhr@em.agr.ca
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't