A Technique Combining Immunoprecipitation and RT‐PCR for RNA Plant Virus Detection

Abstract

Plant virus identification and characterization can be accomplished by several methods involving their morphological, physical, biological, cytological, serological and molecular properties. The use of molecular techniques is increasing worldwide, and some have been developed for identification and characterization of plant viruses. Reverse transcription polymerase chain reaction (RT-PCR) has been shown to be a suitable method for research with RNA plant viruses. In this study, a new approach of RT-PCR involving previous virus immunoprecipitation (IP) was used for RNA amplification of five virus species of the genera Comovirus, Cucumovirus, Potyvirus and Sobemovirus from infected plant tissues. IP-RT-PCR was practical, sensitive and minimized problems with total RNA extractions from infected tissues. The technique provides partial virus particle purification by its specific immunoprecipitation, and it should be especially useful for RNA amplification of viruses that occur in low or variable concentrations in plant tissues or when the tissues contain various forms of RT-PCR amplification inhibitors.

Narcissus yellow stripe virus and Narcissus mosaic virus detection in Narcissus via multiplex TaqMan-based reverse transcription-PCR assay

“…Currently, virus isolates have been detected on seeds, screening propagation materials or other parts mostly via electron microscopy, immunological diagnosis such as enzyme-linked immunosorbent assay, molecular diagnosis such as restriction fragment length polymorphism, thermostable amplification-based methods, isothermal amplification methods, nucleic acid sequence hybridization techniques and next-generation sequencing, with genetic information carried via nucleic acid and microarray (Lima et al 2014;Yadav and Khurana 2016). Consequently, assays based on RT-PCR methods were established to detect Narcissus latent virus (NLV), NYSV and NMV in culture fields (Berniak et al 2013;Hori et al 2013).…”