An improved reverse transcription-polymerase chain reaction (RT-PCR) assay for the detection of two cherry flexiviruses in Prunus spp.

Li, Ruhui; Mock, Ray

doi:10.1016/j.jviromet.2005.05.019

Cited by 26 publications

(10 citation statements)

References 11 publications

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“…In this study, we used primers CGRMV1 and CGRMV2 designed for detection of both CGRMV and CNRMV viruses (Li & Mock 2005). The analysis showed the identity of these primers with the majority of sequences of both CNRMV and CGRMV available from GenBank or NGS sequences (Supplementary file, Figure S1 and S2).…”

Section: Discussionmentioning

confidence: 99%

“…Total RNA was isolated for RT-PCR screening using an RNeasy Plant Mini Kit (QIAGEN, Hilden, Germany) according to the manufacturer's instructions, reverse transcribed using random primers (iScript cDNA Synthesis Kit; Bio-Rad Laboratories, Irvine, USA), then amplified by PCR (TopBio, Prague, Czech Republic) using the primers CGRMV1 (5'-CCT-CATTCACATAGCTTAGGTTT-3') and CGRMV2 (5'-ACTTTAGCTTCGCCCCGTG-3') (Li & Mock 2005) Table S1). All PCR products thus obtained were sequenced (GATC, Constance, Germany) using the Sanger sequencing method.…”

Section: Methodsmentioning

confidence: 99%

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Cherry necrotic rusty mottle and Cherry green ring mottle viruses in Czech cherry germplasm

Špak¹,

Přibylová²,

Šafářová³

et al. 2017

Plant Prot. Sci.

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AbstractŠpak J., Přibylová J., Šafářová D., Lenz O., Koloniuk I., Navrátil M., Fránová J., Špaková V., Paprštein F. (2017) Using reverse transcription polymerase chain reaction, 160 sweet and sour cherry trees from a germplasm collection, orchards, and wild trees in the Czech Republic were screened for the presence of Cherry necrotic rusty mottle virus (CNRMV) and Cherry green ring mottle virus (CGRMV). The viruses were detected exclusively in sweet cherry trees in the germplasm collection, with CNRMV determined in two trees and CGRMV in four trees. Using next-generation sequencing, nearly complete genomic sequences (complete ORFs) were obtained for one CNRMV and three CGRMV isolates. Their relatedness to GenBank sequences of isolates from different countries together with negative results from screening outside of the germplasm collection suggests that the viruses had been imported with accessions.

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Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Cherry necrotic rusty mottle and Cherry green ring mottle viruses in Czech cherry germplasm

Špak¹,

Přibylová²,

Šafářová³

et al. 2017

Plant Prot. Sci.

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“…alternative tools for specific diagnosis in quarantine and certification programmes (Henson and French 1993;Li and Mock 2005).…”

Section: Archives Of Phytopathology and Plant Protection 1011mentioning

confidence: 99%

Partial characterisation of a cherry isolate of a newly emerging stone fruit virus:Plum bark necrosis stem pitting associated virus

Sipahioğlu

Usta

Oksuz

2011

Archives Of Phytopathology And Plant Protection

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Plum bark necrosis stem pitting associated virus (PBNSPaV) has occurred globally over the last two decades and is becoming one of the major agricultural issues of stone fruits. The virus was detected for the first time in plums and has been reported in the other stone fruit species. The agent, whose dissemination mode is still unknown, has been first reported in cherries in Turkey grown in Malatya province. In this study, the K1 isolate of PBNSPaV, identified in a sweet cherry plant with severe stem pitting and gumming symptoms on its trunk, was partially characterised. A fragment of Hsp70h gene located on ORF3 of viral genome has been cloned, sequenced and analysed phylogenetically (Accession number. FJ231498). The PBNSPaV-K1 isolate showed 93-96% nucleotide sequence identity to sequences of Italian and American isolates in databases. An RNA probe has been raised for fast and reliable detection of the agent by molecular hybridisation. The studies on development of genome specific primers for the detection of the isolate by one-tube RT-PCR have failed.

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“…Primers should have an annealing temperature (T m ) of ∼60 • C and lack sequences with self complementarity. Primer pairs that meet these criteria may be compared using RT-PCR to detect the viruses of interest (Li and Mock, 2005). The consensus primers used in the protocol are based on two highly conserved regions adjacent to the coat protein (CP) genes and have been used successfully to detect all CGRMV and CNRMV isolates tested.…”

Section: Rt-pcr Assay For Identification Of Viral Pathogensmentioning

confidence: 99%

Reverse Transcription‐Polymerase Chain Reaction‐Based Detection of Plant Viruses

Hartung

2007

CP Microbiology

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A one-step reverse transcription-polymerase chain reaction (RT-PCR) is used to detect two cherry flexiviruses, Cherry green ring mottle virus (CGRMV) and Cherry necrotic rusty mottle virus (CNRMV), in Prunus species. This unit presents procedures for collection of plant samples, preparation of total nucleic acids, viral RNA-rich or total RNA extracts from plant tissues, and subsequent amplification of the viral targets by one-step RT-PCR using a pair of consensus primers. The PCR amplicons are visualized by electrophoresis in a 1% agarose gel containing ethidium bromide in TAE buffer and viewed under ultraviolet light. This procedure is rapid, sensitive, reliable, and cost-effective and is generally useful on a wide variety of plant/virus systems. The use of a semi-automatic homogenizer for sample preparation and one-tube RT-PCR for virus detection makes this approach ideal for screening large numbers of samples. Curr. Protoc. Microbiol. 6:16C.

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An improved reverse transcription-polymerase chain reaction (RT-PCR) assay for the detection of two cherry flexiviruses in Prunus spp.

Cited by 26 publications

References 11 publications

Cherry necrotic rusty mottle and Cherry green ring mottle viruses in Czech cherry germplasm

Cherry necrotic rusty mottle and Cherry green ring mottle viruses in Czech cherry germplasm

Partial characterisation of a cherry isolate of a newly emerging stone fruit virus:Plum bark necrosis stem pitting associated virus

Reverse Transcription‐Polymerase Chain Reaction‐Based Detection of Plant Viruses

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