A PCR membrane spot assay for the detection of plum pox virus RNA in bark of infected trees
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2019, Journal of Virological MethodsCitation Excerpt :Nevertheless, the lack of obvious symptoms of disease caused by LChV-1 hampers efficient diagnosis. Moreover, other evident reasons are making detection a challenging process: (i) in RT-PCR, plant extracts contaminants from Prunus frequently affect RNA quality and inhibition of subsequent tests (Candresse et al., 1998); (ii) in all these tests, high variability in virus concentrations interferes with the analytical sensitivity of the test, making LChV-1 detection fastidious (Korschineck et al., 1991; Demeke and Adams, 1992; Nassuth et al., 2000; Li et al., 2013); (iii) in molecular tests, inclusiveness is hindered by the relatively high level of variability between isolates of LChV-1. A fast, cheap and reliable detection technique for LChV-1 is therefore essential.
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2018, Analytical BiochemistryCitation Excerpt :The nucleic acid extraction represents approximately one-half of the cost of assay. Moreover, viral DNA or RNA is frequently recovered carrying plant inhibitor compounds that may interfere with PCR and RT-PCR [41–48], all of which can be seen as a disadvantage. In contrast RPA bring into diagnostics the possibility of utilizing crude sap extracted directly from plant tissues, which improve the assay cost-effectiveness.
A rapid silica spin column-based method of RNA extraction from fruit trees for RT-PCR detection of viruses
2017, Journal of Virological MethodsCitation Excerpt :Virus detection is necessary for the production of certified virus-free propagation materials of fruit trees. The leaves and/or bark tissues of most fruit trees contain some amount of polysaccharides and polyphenolic compounds (Demeke and Adams, 1992; Gambino et al., 2008), which usually are difficult to be removed and inhibit reverse transcription and PCR reactions (Asif et al., 2000; Demeke and Adams, 1992; Jones and McGavin, 2002; Korschineck et al., 1991; Li et al., 2008; Loomis, 1974; Nassuth et al., 2000; Pandey et al., 1996; Salzman et al., 1999; Staub et al., 1995). Moreover, fruit tree viruses usually occur at low titers and with uneven distribution in infected plants, their detection is difficult, which is also a challenge for high-quality RNA recovery.
Rapid and sensitive detection of Little cherry virus 2 using isothermal reverse transcription-recombinase polymerase amplification
2014, Journal of Virological MethodsCitation Excerpt :Currently, the most commonly used detection method is RT-PCR (Vitushkina et al., 1997). Viral RNA can be reliably detected in nucleic acids recovered from plant tissues, but contaminants from plant extracts frequently inhibit PCR (Korschineck et al., 1991; Demeke and Adams, 1992; Henson and French, 1993; Staub et al., 1995; Nassuth et al., 2000; Jones and McGavin, 2002; Tairo et al., 2006; Li et al., 2008). Current RT-PCR methods require careful preparation of target nucleic acids from the sample, especially from fruit trees, and in our laboratory, RNA isolation represents approximately one-half the cost of analysis.
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2008, Journal of Virological Methods