A novel multiplex RT-PCR probe capture hybridization (RT-PCR-ELISA) for simultaneous detection of six viroids in four genera: Apscaviroid, Hostuviroid, Pelamoviroid, and Pospiviroid

Shamloul, A. M.; Faggioli, F.; Keith, Jerry M.; Hadidi, A.

doi:10.1016/s0166-0934(02)00090-3

Cited by 47 publications

(25 citation statements)

References 15 publications

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“…The use of multiplex RT-PCR for the simultaneous detection of more than two viruses or viroids using a mixture of specific primer pairs (Di Serio et al, 2002;Ito et al, 2002;Levy et al, 1992;Nie and Singh, 2001;Shamloul et al, 2002) has been described. Multiplex RT-PCR has the potential to dramatically reduce the diagnostic cost by reducing the number of assays to be performed if the detection of several viroids is needed (Hadidi and Candresse, 2003).…”

Section: Discussionmentioning

confidence: 99%

“…The multiplex PCR or RT-PCR method has been established for the simultaneous detection of several viruses (Bariana et al, 1994;Jacobi et al, 1998;Singh et al, 1996) and viroids (Di Serio et al, 2002;Ito et al, 2002;Levy et al, 1992;Nie and Singh, 2001;Shamloul et al, 2002). The multiplex PCR method reduces the time and cost required for template preparation for virus-or viroidfree plant screening.…”

Section: Introductionmentioning

confidence: 99%

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A Simple and Simultaneous Detection Method for Two Different Viroids Infecting Chrysanthemum by Multiplex Direct RT-PCR

Hosokawa¹,

Shiba²,

Kawabe³

et al. 2007

J. Japan. Soc. Hort. Sci.

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The economically important Chrysanthemum stunt viroid (CSVd) and Chrysanthemum chlorotic mottle viroid (CChMVd) were detected from infected chrysanthemum (Dendranthema grandiflorum) plants by a multiplex reverse transcription-polymerase chain reaction (RT-PCR) assay developed in this investigation. The antisense hexamer AAAGGA (5'-3') was designed. The antisense hexamer 5'AAAGGA 3' annealing at 5'TCCTTT3' is located at nucleotide positions 186-191 and 245-250 of CSVd (gb: AB006737), and 231-236 of CChMVd in their sequences. Using CSVd-and CChMVd-cDNA templates which were transcribed simultaneously by the hexamer, the following multiplex PCR could detect both viroids from doubly-infected plants without nonspecific amplification. When the hexamer was used for the RT reaction, the sensitivity of detection of CSVd or CChMVd by multiplex RT-PCR was similar to that of standard RT-PCR when each viroid was detected separately. Furthermore, multiplex RT-PCR successfully detected both CSVd and CChMVd in direct templates obtained by inserting a syringe needle into the stem, leaf, and shoot tips of infected chrysanthemum plants. The direct multiplex RT-PCR method developed in this study may reduce the cost, time, and labor required for the production of viroid-free chrysanthemum plants.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

A Simple and Simultaneous Detection Method for Two Different Viroids Infecting Chrysanthemum by Multiplex Direct RT-PCR

Hosokawa¹,

Shiba²,

Kawabe³

et al. 2007

J. Japan. Soc. Hort. Sci.

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“…For the detection of HSVd and Citrus exocortis viroid (CEVd), the methods of Shamloul et al (2002) and Gross et al (1982), respectively, were used. Additionally, two primer sets were designed to specifically detect the full-length genome of GYSPVd-1 [cGYSVd1: 5′-CGAGGCTCACTCCCCCTCTGCC-3′ / h-GYSVd1: 5′-TCGTCGACGAAGGGGTGCACTCC-3′], and -2 [c-GYSVd2: 5′-GGTCCGCGGAGGCCTTCCGAGG-3′ / h-GYSVd2: 5′-TGCAGAGAAAAGAAGAAGGGCCCAG-3′] (Tangkanchanapas et al, 2015).…”

Section: Figurementioning

confidence: 99%

First report of Grapevine yellow speckle viroid‐2 infecting grapevine (Vitis vinifera) in Thailand

Tangkanchanapas¹,

Reanwarakorn²,

Juenak³

et al. 2017

New Disease Reports

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“…This diagnostic approach is resource intensive, and the results are greatly affected by environmental conditions. Nucleic acid-based detection techniques such as dot blot hybridization probes and reverse transcription polymerase chain reaction (RT-PCR) have been developed to detect pome viroids (Faggioli et al 2001;Shamloul et al 2002;Lolic et al 2007;Lin et al 2011). Simultaneous detection of ADFVd, ASSVd and PBCVd using a single pair of primers was described (Faggioli et al 2001).…”

Section: Introductionmentioning

confidence: 99%

One-step multiplex RT-PCR for simultaneous detection of four pome tree viroids

Lin

Li²,

Mock³

et al. 2012

Eur J Plant Pathol

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Apple scar skin viroid (ASSVd), Apple dimple fruit viroid (ADFVd), Apple fruit crinkle viroid (AFCVd), and Pear blister canker viroid (PBCVd) infect pome fruit trees. These viroids are important quarantine pathogens for the international movement of pome germplasm. A single-step multiplex reverse transcription polymerase chain reaction assay (mRT-PCR) was developed for the simultaneous detection of these viroids. Four pairs of primers specific for each of the four viroids were used to amplify PCR products of different sizes that can be resolved by agarose gel electrophoresis. Amplification of a plant 18S rRNA was included in the assay as an internal control. Amplicons of 371 bp (AFCVd), 270 bp (ADFVd), 186 bp (ASSVd), 120 bp (PBCVd), and 844 bp (18S rRNA) were obtained in both uniplex and mRT-PCR assays. The identities of the amplification products were confirmed by sequencing. The specificities and limits of detection for all four viroids by uniplex and mRT-PCR assays were comparable. The assay was further validated using samples from pome trees inoculated with all four viroids, as well as field samples from commercial orchards in Colorado. All four viroids were detected from inoculated pear trees and up to three viroids were detected from inoculated apple trees. This is a simple, rapid and cost-effective technique to detect these four viroids in fruit trees. The procedure is especially applicable to certification and quarantine programs, where numerous samples must be tested for all four viroids.Keywords Apscaviroid . Apple dimple fruit viroid . Apple fruit crinkle viroid . Apple scar skin viroid . Multiplex RT-PCR . Pear blister canker viroid

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A novel multiplex RT-PCR probe capture hybridization (RT-PCR-ELISA) for simultaneous detection of six viroids in four genera: Apscaviroid, Hostuviroid, Pelamoviroid, and Pospiviroid

Cited by 47 publications

References 15 publications

A Simple and Simultaneous Detection Method for Two Different Viroids Infecting Chrysanthemum by Multiplex Direct RT-PCR

A Simple and Simultaneous Detection Method for Two Different Viroids Infecting Chrysanthemum by Multiplex Direct RT-PCR

First report of Grapevine yellow speckle viroid‐2 infecting grapevine (Vitis vinifera) in Thailand

One-step multiplex RT-PCR for simultaneous detection of four pome tree viroids

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