Multimeric non-radioactive cRNA probes improve detection of potato spindle tuber viroid (PSTVd)

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

“…Regarding sensitivity, the nearly full-length probe described here performs as well as previous radioactive or multimeric probes described for other viroids (Roy et al, 1989;Candresse et al, 1990;Kanematsu et al, 1991;Singh et al, 1994), clearly detecting subpicogram amounts of HSVd RNAs. HSVd was also detected in dilutions of infected plant extracts corresponding to 12.5 ng of analysed tissue (Figure lb).…”

“…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.…”

RT-PCR analysis of PSTVd aphid transmission in association with PLRV

“…Following electrophoresis, gels were denatured and neutralised and DNA was transferred to nitrocellulose BA-S-83 (Schleicher and Schuell, Keene, NH) in a Vacugene XL apparatus (Pharmacia LKB Biotechnology) (Singh et al 1995). A cDNA tetramer clone of PSTVd labelled with digoxigenin (Singh et al 1994) was used as a probe to verify the PCR band specificity to PSTVd.…”

“…Isolation of nucleic acids from leaves and aphids was carried out as described (Singh et al 1995). Presence of PLRV and PSTVd was confirmed in plants by the enzyme-linked immunosorbent assay (Singh et al 1996b) and nucleic acid hybridization (Singh et al 1994) respectively.…”

Analysis of immunogenicity of minor envelope protein GP3 of porcine reproductive and respiratory syndrome virus in mice

“…Viroid detection mainly depends either on time-consuming biological assays or on molecular techniques aimed to detect the presence of their RNA genomes [3]. Commonly applied techniques are the direct visualization of viroid RNAs after gel electrophoresis [10,11], molecular hybridization [12][13][14][15][16][17][18][19] and PCR amplification [20][21][22][23][24][25][26][27][28]. The choice and applicability of these methods for different viroids depends on the diagnostic purposes, the availability of trained personnel and viroid titer in infected tissues.…”