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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » National Germplasm Resources Laboratory » Research » Publications at this Location » Publication #328095

Research Project: CHARACTERIZING, DETECTING, AND ELIMINATING PATHOGENS TO ENABLE THE SAFE INTRODUCTION OF PLANT GENETIC RESOURCES

Location: National Germplasm Resources Laboratory

Title: Simultaneous detection and differentiation of three Potyviridae viruses by a multiplex TaqMan real time RT-PCR assay

Author
item LAN, PINGXIU - Yunnan Agricultural University
item ABAD, JORGE - Animal And Plant Health Inspection Service (APHIS)
item PU, LINGLING - South China Agricultural University
item LI, FAN - Yunnan Agricultural University
item Li, Ruhui

Submitted to: Journal of Virological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/12/2017
Publication Date: 9/17/2017
Citation: Lan, P., Abad, J., Pu, L., Li, F., Li, R. 2017. Simultaneous detection and differentiation of three Potyviridae viruses by a multiplex TaqMan real time RT-PCR assay. Journal of Virological Methods. https://doi:10.1016/j.jviromet.2017.09.006.

Interpretive Summary: Sweet potato is one of most important root/tuber crops in the world. More than 30 different viruses infect and cause disease in this crop. Sweet potato viral disease caused by mixed infections of several viruses, including Sweet potato virus G, Sweet potato latent virus (SPLV) and Sweet potato mild mottle virus (SPMMV), is the most devastating viral disease worldwide and can reduce the yields up to 80-90%. In this study, a simple, sensitive, and cost-effective method is developed to detect these three viruses in one tube. The assay is reliable for detecting the viruses using samples from the greenhouse and from sweet potato production fields. The assay should be very useful in quarantine and certification programs, and as a tool to facilitate field surveys and other diagnostics that support disease management efforts.

Technical Abstract: A multiplex TaqMan real time RT-PCR was developed for detection and differentiation of Sweet potato virus G, Sweet potato latent virus and Sweet potato mild mottle virus in one tube. Amplification and detection of a fluorogenic cytochrome oxidase gene was included as an internal control. The assay was compared with a multiplex RT-PCR developed in the initial study for the detection and differentiation of the three viruses and host 18S rRNA. Primers and/or probes of the two assays were designed from conserved regions of each virus. The two assays were optimized for primers/probes and primer concentrations and thermal cycling conditions. Sensitivity and specificity of the assays were compared each other and with other assay. Both assays were evaluated by 74 field samples original from five different provinces of China. Results showed that the TaqMan real time RT-PCR offered rapid, sensitive, effective and reliable for the simultaneous detection and differentiation of the three viruses in sweet potato plants. The assay will be useful to quarantine and certification programs and virus surveys when large numbers of samples are tested.