Use of a<sup>32</sup>P-Labeled Transcribed RNA Probe for Dot Hybridization Detection of Plum Pox Virus

Varveri, C.; Candresse, Thierry; Cugusi, M.; Ravelonandro, Michel; Dunez, Jean

doi:10.1094/phyto-78-1280

Cited by 46 publications

(31 citation statements)

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“…In an attempt to overcome this problem RT-PCR using immunocaptured viral particles (IC-RT-PCR) or total RNA as input and nested RT-PCR have been used for detection of PPV (Wetzel et al 1991;Candresse et al 1995;Adams et al 1999) and PNRSV (Spiegel et al 1996;Rosner et al 1998;Helguera et al 2001). Furthermore, methods based on RNA hybridization using radioactive (Varveri et al 1988;Crosslin et al 1992) and non-radioactive (Heuss-La Rosa et al 1995;Heuss et al 1999) cRNA probes have been employed for their detection.…”

Section: Virus Detectionmentioning

confidence: 99%

Elimination of PPV and PNRSV through thermotherapy and meristem-tip culture in nectarine

et al. 2003

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The plum pox virus (PPV) and prunus necrotic ringspot virus (PNRSV) cause serious disease problems in stone-fruit trees. In this work, the possibility of obtaining plant material free from these viruses through thermotherapy and meristem-tip culture from infected nectarine shoots (Prunus persica var. nectarina Max, cv. 'Arm King') was studied. In addition, the detection of these viruses in in vitro cultures and young acclimatized plantlets with double antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) and multiplex reverse transcriptase-polymerase chain reaction (RT-PCR) was studied. Meristem-tip explants (0.8-1.3 mm) derived from sprouted buds of winter wood and spring shoots from field grown plants had a 2-5% regeneration response. However, application of thermotherapy to potted nectarine trees (3 weeks at a maximum temperature of 35 degrees C) facilitated excision of longer meristem tips (1.3-2.0 mm) that resulted in a significantly higher regeneration response (38%) in woody plant medium (WPM) without plant growth regulators. Such explants formed multiple shoots with the addition of 8 microM benzylaminopurine and 0.8 microM indoleacetic acid. When they were tested for the presence of PPV and PNRSV, 86% and 81% were found to be virus-free as detected by DAS-ELISA and multiplex RT-PCR, respectively. Individual shoots excised from virus-free cultures readily rooted in vitro (half-strength WPM plus 2 microM indolebutyric acid) and grew to plantlets. The combination of an efficient protocol for virus elimination and the establishment of highly sensitive diagnostics resulted in the production of nectarine plants free from PPV and PNRSV.

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Section: Virus Detectionmentioning

confidence: 99%

Elimination of PPV and PNRSV through thermotherapy and meristem-tip culture in nectarine

et al. 2003

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“…The digoxigenin-11-dUTP-labelled cDNA probes, corresponding to BCMV structural and non-structural genes (Varveri et al 1988;Wetzel et al 1991), were amplified by 1434 M.A. El-Sawy et al…”

Section: Probe Preparationmentioning

confidence: 99%

Serological and molecular characterisations of the Egyptian isolate ofBean common mosaic virus

El-Sawy

Mohamed

Elsharkawy

2013

Archives Of Phytopathology And Plant Protection

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Bean common mosaic virus (BCMV) was isolated from the naturally infected bean plants collected from the Kafr El-Sheikh and El-Gharbia Governorates. BCMV induced sever mosaic, vein banding, malformation, leaf curling and stunting on bean plants cv. Giza 6. The isolated virus was propagated in bean plants cv. Giza 6. The identification of BCMV was carried out serologically by an indirect enzyme-linked immunosorbent assay using BCMV antiserum. Positive reaction indicated that the virus under study was related serologically to Potyvirus. The molecular biology techniques were used to identify and characterise the coat protein gene of BCMV. Oligonucleotide primers were designed for BCMV according to the published nucleotide sequences of BCMV and were successfully amplified with a DNA fragment (300 bp) from BCMV CP gene by RT-PCR. The total RNA was extracted from bean leaves and was reverse-transcribed and amplified using the oligonucleotide primer. The amplified product was analysed by gel electrophoresis. Also, Southern and dot blot hybridisations were used to establish the authenticity and specificity to the RT-PCRamplified products of BCMV. The nucleotide sequences of the Egyptian isolate of BCMV/CP showed similarity with an isolate (BCMV-NY 15) which belongs to Puerto Rico.

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“…This approach has been used to detect potato spindle tuber viroid (86,106), barley yellow dwarf virus (42), X potato virus (9), avocado sunblotch viroid (8), plum pox virus (118), and citrus exocortis viroid (33).…”

Section: Double-stranded Dn a Probesmentioning

confidence: 99%

“…RNA probes also have less background than DNA probes (3). Although ssDNA probes and RNA probes have a higher sensitivity than dsDNA probes, ssDNA probes (49) and RNA probes (63,118) have not been frequently used to detect environmentally relevant organisms (Table 1). Perhaps they have not been exploited because of the extra cloning steps required to make RNA and ssDNA probes and the extra caution needed in handling RNA probes.…”

Section: Single-str Anded Dn a And Rn A Probesmentioning

confidence: 99%

Environmental Application of Nucleic Acid Hybridization

Sayler

Layton

1990

Annu. Rev. Microbiol.

174

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Use of a³²P-Labeled Transcribed RNA Probe for Dot Hybridization Detection of Plum Pox Virus

Cited by 46 publications

References 0 publications

Elimination of PPV and PNRSV through thermotherapy and meristem-tip culture in nectarine

Elimination of PPV and PNRSV through thermotherapy and meristem-tip culture in nectarine

Serological and molecular characterisations of the Egyptian isolate ofBean common mosaic virus

Environmental Application of Nucleic Acid Hybridization

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Use of a32P-Labeled Transcribed RNA Probe for Dot Hybridization Detection of Plum Pox Virus

Cited by 46 publications

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Elimination of PPV and PNRSV through thermotherapy and meristem-tip culture in nectarine

Elimination of PPV and PNRSV through thermotherapy and meristem-tip culture in nectarine

Serological and molecular characterisations of the Egyptian isolate ofBean common mosaic virus

Environmental Application of Nucleic Acid Hybridization

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Use of a³²P-Labeled Transcribed RNA Probe for Dot Hybridization Detection of Plum Pox Virus