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An improved reverse transcription-polymerase chain reaction (RT-PCR) assay for the detection of two cherry flexiviruses in Prunus spp
- Source :
- Journal of Virological Methods. 129:162-169
- Publication Year :
- 2005
- Publisher :
- Elsevier BV, 2005.
-
Abstract
- A one-step reverse transcription-polymerase chain reaction (RT-PCR) procedure was developed to detect Cherry green ring mottle virus (CGRMV) and Cherry necrotic rusty mottle virus (CNRMV) in woody indicators and naturally infected Prunus spp. Viral RNA suitable for RT-PCR was obtained by a simple trapping method that did not require either extraction of double-stranded RNA (dsRNA) or total RNA, availability of virus antibodies, or purification of viral particles. Consensus primers, degenerate primers and virus-specific primers, whose designs were based on alignments of available cherry flexivirus sequences, were tested to amplify viral genomic fragments of six CGRMV isolates and one CNRMV isolate. RT-PCR allowed CGRMV detection in total RNA and viral RNA preparations equivalent to 400mug and 4mug of infected leaf tissue, respectively. CGRMV was detected in tender shoots, leaves, bark and root tips, and the strongest bands were obtained using young leaves. Detection was less consistent in summer when the temperature was elevated and plant tissues were old. A direct comparison of the RT-PCR and grafting assays indicated that the RT-PCR assay is sensitive, rapid and reliable. The method will improve the routine diagnosis of cherry flexiviruses in Prunus spp.
- Subjects :
- Reverse Transcriptase Polymerase Chain Reaction
Molecular Sequence Data
RNA
Biology
Plant Roots
Sensitivity and Specificity
Virology
Virus
Plant Viruses
law.invention
Plant Leaves
Reverse transcription polymerase chain reaction
Prunus
RNA silencing
Real-time polymerase chain reaction
Species Specificity
law
Plant virus
Capsid Proteins
Plant Shoots
Polymerase chain reaction
DNA Primers
Plant Diseases
Subjects
Details
- ISSN :
- 01660934
- Volume :
- 129
- Database :
- OpenAIRE
- Journal :
- Journal of Virological Methods
- Accession number :
- edsair.doi.dedup.....30149768ebf3e148426124bc1c752c2f
- Full Text :
- https://doi.org/10.1016/j.jviromet.2005.05.019