Abstract
BackgroundWheat yellow mosaic virus (WYMV) is an important pathogen in China and other countries. It is the member of the genus Bymovirus and transmitted primarily by Polymyxa graminis. The incidence of wheat infections in endemic areas has risen in recent years. Prompt and dependable identification of WYMV is a critical component of response to suspect cases.MethodsIn this study, a one step real-time RT-PCR, followed by standard curve analysis for the detection and identification of WYMV, was developed. Two reference genes, 18s RNA and β-actin were selected in order to adjust the veracity of the real-time RT-PCR assay.ResultsWe developed a one-step Taqman-based real-time quantitative RT-PCR (RT-qPCR) assay targeting the conserved region of the 879 bp long full-length WYMV coat protein gene. The accuracy of normalized data was analyzed along with appropriate internal control genes: β-actin and 18s rRNA which were included in detecting of WYMV-infected wheat leaf tissues. The detectable end point sensitivity in RT-qPCR assay was reaching the minimum limit of the quantitative assay and the measurable copy numbers were about 30 at106-fold dilution of total RNA. This value was close to 104-fold more sensitive than that of indirect enzyme-linked immunosorbent assay. More positive samples were detected by RT-qPCR assay than gel-based RT-PCR when detecting the suspected samples collected from 8 regions of China. Based on presented results, RT-qPCR will provide a valuable method for the quantitative detection of WYMV.ConclusionsThe Taqman-based RT-qPCR assay is a faster, simpler, more sensitive and less expensive procedure for detection and quantification of WYMV than other currently used methods.
Highlights
Wheat yellow mosaic virus (WYMV) is an important pathogen in China and other countries
Filamentous viruses transmitted by P. graminis cause similar diseases of wheat in Europe, Asia and North America [7,8]. These viruses have been described as Wheat spindle streak mosaic virus (WSSMV) and WYMV in different countries [7,9,10]
A one step real-time quantitative RT-PCR (RT-qPCR) assay was established for the detection, discrimination, and quantification of WYMV in wheat plants
Summary
Wheat yellow mosaic virus (WYMV) is an important pathogen in China and other countries. It is the member of the genus Bymovirus and transmitted primarily by Polymyxa graminis. Filamentous viruses transmitted by P. graminis cause similar diseases of wheat in Europe, Asia and North America [7,8]. These viruses have been described as Wheat spindle streak mosaic virus (WSSMV) and WYMV in different countries [7,9,10]. The virus causing wheat soil-borne mosaic diseases has been identified as WYMV in most areas of China [11,12]. RNA1 encodes for the coat protein (CP) and six others: P3, 7 KDa, nuclear inclusion protein a (NIa), nuclear inclusion protein b (NIb), cytoplasmic inclusion protein (CI) and 14 KDa; RNA2 encodes for a polyprotein that contains 28- and 72-kDa proteins [13,14]
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