Highly sensitive digoxigenin-labelled DNA probe for the detection of potato spindle tuber viroid

Welnicki, M.; Hiruki, C.

doi:10.1016/0166-0934(92)90128-z

Cited by 22 publications

(7 citation statements)

References 16 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Earlier, NASH techniques based on radioactive and non radioactive probes (Fluorescein labelled probes) were used to detect PSTVd which had a detection limit of 10 pg and 10 ng, respectively (Verma et al 2006). In case of purified PSTVd RNA, sensitivity of digoxigenin labelled probe was up to 2.5 pg (Welnicki and Hiruki 1992). But in this study, using realtime RT-PCR assay we could detect up to 0.025 fg total RNA, which indicates that the developed realtime RT-PCR highly sensitive than NASH assay.…”

Section: Discussioncontrasting

confidence: 62%

Duplex realtime RT-PCR assay for the detection of Potato spindle tuber viroid (PSTVd) along with ef 1-α gene of potato

et al. 2015

View full text Add to dashboard Cite

SYBR green based realtime RT-PCR assay coupled with melt curve analysis was developed for the detection of Potato spindle tuber viroid (PSTVd) along with and without internal control from potato. The amplification of the specific targets was identified by their melting points viz., 85.93±0.22, 85.62±0.34 and 82.07 ±0.23 for primer pairs PSTVd-1F/PSTVd-1R, PSTVd-NFP/PSTVd-NRP and PSTVd-QFP1/PSTVd-QRP1, respectively. The realtime RT-PCR assay was 1×10 4 times more sensitive than RT-PCR assay and the assay could detect up to 20 copies of the target using serially diluted plasmid and up to 0.025 fg of total RNA from infected tissues diluted in healthy plant RNA. Duplex realtime RT-PCR assay was also standardized to detect PSTVd along with elongation factor 1-α (ef-1α) gene, a stable housekeeping gene of potato. Duplex realtime RT-PCR assay showed two melt peaks indicating the successful amplification both the PSTVd RNA and internal control RNA. The developed assays could consistently detect PSTVd and proved to be highly sensitive and rapid for the detection of PSTVd in post entry quarantine testing.

show abstract

Section: Discussioncontrasting

confidence: 62%

Duplex realtime RT-PCR assay for the detection of Potato spindle tuber viroid (PSTVd) along with ef 1-α gene of potato

et al. 2015

View full text Add to dashboard Cite

show abstract

“…A further improvement arose with the advent of non-radioactive methods for the detection of hybridized probes, allowing their progressive introduction as tools in the study of plant viruses (Habili et al, 1987;Eweida etal., 1990;Fouly et al, 1992;Gemmrich etal., 1993;M~s et al, 1993;Dietzgen etal., 1994;Harper and Creamer, 1995;S~nchez-Navarro et al, 1996). Several reports have already described the use of these non-radioactive probes to detect viroid RNAs, albeit most of them correspond to potato spindle tuber viroid (PSTVd) (McInnes et al, 1989;Roy et al, 1989;Candresse et al, 1990;Hopp et al, 1991;Kanematsu et al, 1991;Welnicki and Hiruki, 1992;Podleckis et al, 1993;Singh et al, 1994). Recently, a non-radioactive tissue-print hybridization method was reported for viroid detection (Romero-Durb~n et al, 1995), although the authors encountered problems in the detection of HSVd.…”

Section: Introductionmentioning

confidence: 99%

Studies on the diagnosis of hop stunt viroid in fruit trees: Identification of new hosts and application of a nucleic acid extraction procedure based on non-organic solvents

et al. 1996

View full text Add to dashboard Cite

A non-radioactive digoxigenin-labelled RNA probe specific for hop stunt viroid (HSVd) diagnosis has been developed. The high sensitivity and specificity of this RNA probe in dot blot hybridizations to nucleic acids from field samples, allowed the confirmation of the presence of HSVd in apricot (Prunus armeniaca L.) and its detection in two fruit tree species not previously described as hosts of this pathogen, almond (Prunus dulcis Miller) and pomegranate (Punica granatum L.). This result supports and extends the notion of the world wide distribution of HSVd, infecting cultivated fruit trees. HSVd was also found to accumulate to much higher levels in mature apricot fruits than in leaves. Additionally, a sample processing procedure which does not involve the use of organic solvents was demonstrated to render faithful results when used for viroid detection. The combined reliability and facility of use of both this extraction procedure and the non-radioactive probe will benefit agronomic investigations addressing the detection and eradication of HSVd. Other applications of the work described here, as the study of possible causal relations between specific disorders and HSVd infection, are also discussed.

show abstract

“…Owens & Diener, 1981;Maule et al, 1983;Garger et al, 1983;Hull, 1984Hull, , 1993Baulcombe et al, 1984;Sela et al, 1984;Varveri et al, 1987) particularly since the use of non-radioactive precursor to label the nucleic acid probes made this technique more accessible (e.g. Habili et al, 1987;Roy et al, 1988;Eweida et al, 1990;Welnicki & Hiruki, 1992;Fouly et al, 1992;Más et al, 1993;Gemmrich et al, 1993;Dietzgen et al, 1994;Más & Pallás, 1995).…”

Section: Introductionmentioning

confidence: 99%

Non‐radioactive molecular hybridization detection of carnation mottle virus in infected carnations and its comparison to serological and biological techniques

Sánchez‐Navarro

Cano²,

Pallás

1996

Plant Pathology

View full text Add to dashboard Cite

A non-radioactive molecular hybridization method was developed to detect carnation mottle virus (CarMV) in carnation plants. This method was compared to the standard ELISA test and biological assays. For crude plant extracts, the molecular hybridization technique proved to be at least 125 times more sensitive than double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA). The method was adapted to avoid the necessity to clarify the samples by centrifugation and to use pipettes to load them. By introducing this modification, the hybridization assay was found to be a suitable method for routine virus detection. The relative benefits of using this procedure for large-scale indexing are discussed.

show abstract

Highly sensitive digoxigenin-labelled DNA probe for the detection of potato spindle tuber viroid

Cited by 22 publications

References 16 publications

Duplex realtime RT-PCR assay for the detection of Potato spindle tuber viroid (PSTVd) along with ef 1-α gene of potato

Duplex realtime RT-PCR assay for the detection of Potato spindle tuber viroid (PSTVd) along with ef 1-α gene of potato

Studies on the diagnosis of hop stunt viroid in fruit trees: Identification of new hosts and application of a nucleic acid extraction procedure based on non-organic solvents

Non‐radioactive molecular hybridization detection of carnation mottle virus in infected carnations and its comparison to serological and biological techniques

Contact Info

Product

Resources

About