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Crude extract from a healthy potato herb was used as a negative control (CK?). and 14 of them were found to be positive. This indicates that PVS is now Olodaterol prevalent in potato fields in Yunnan Province. Keywords: Potato computer virus S, Tissue print-enzyme-linked immunosorbent assay (ELISA), Dot-ELISA, Double-antibody sandwich (DAS)-ELISA 1.?Introduction Potato is the fourth most important food crop after wheat, rice, and corn in the world, and has high nutritional value. Viral diseases, including Potato computer virus S (PVS), often cause huge losses in potato production. PVS was first found in a potato field in the Netherlands in 1948 and is now in all potato-growing areas worldwide. It is a member of the genus in the family Betaflexiviridae (Lin et al., 2014). The PVS virion is filamentous with 610C710 nm in length and 10C15 nm in diameter, and its genome is single-stranded, positive-sense RNA approximately 8.4 to 8.5 kb long (Foster et al., 1992; Matousek et al., 2005) containing a 5′ cap structure, a 3′ poly(A) tail, and six open reading frames (ORFs). ORF1 encodes a 223-kDa protein containing NTP-binding helicase (HEL), methyltransferase (MTR), and RNA-dependent RNA Olodaterol polymerase (RdRp) domains. ORF2, ORF3, and ORF4 constitute the triple gene block (TGB). ORF2 encodes a protein of 25 kDa with an NTPase/helicase domain. ORF3 and ORF4 encode 12 and 7 kDa proteins, respectively, which is known to be involved in PVS intracellular movement (Mackenzie et al., 1989; Morozov and Solovyev, 2003). ORF5 encodes a 34-kDa coat protein (CP) and ORF6 encodes an 11-kDa nucleotide-binding protein (Lin et al., 2014). PVS only infects members in the Solanaceae and Cheonopodiaceae families (Morelli and Vayda, 1996), with spread between crops being transmitted by aphids (Cerovska and Filigarova, 1995). Spread of the disease between distant regions is thought GDF5 to be caused by trading of PVS-infected seed tubers. Most PVS-infected potato plants do not show obvious symptoms or only mild leaf rugosity, vein deepening and leaf bronzing (Yardimci et al., 2015), but a few highly susceptible cultivars, such as Pentland Javelin and Epicure, can show severe symptoms (Salari et al., 2011). In the field, PVS often co-infects plants along with other viruses such as Potato virus X (PVX) and Potato virus M (PVM), resulting in a 20%C30% reduction of potato yield (Wu et al., 2002). The current PVS management strategy includes breeding disease-resistant varieties and planting virus-free seed tubers. As PVS does not cause clear symptoms in most infected potato plants, establishment of rapid and reliable detection methods is crucial for Olodaterol PVS-resistant potato breeding and PVS-free seed tuber production. Several PVS detection methods have been reported, including assays using indicator plants (Mao, 2009), a reverse transcription-polymerase chain reaction (RT-PCR) (Cheng et al., 2010), and serological detection using PVS-specific polyclonal (Yardimci et al., 2015) and monoclonal (Cerovska and Filigarova, 1995) antibodies (PAbs and MAbs). All, however, have drawbacks: detection using indicator plants takes multiple days to complete, needs greenhouse or growth chamber space, and is questionable in its ability to detect low levels of PVS; RT-PCR is a sensitive and specific assay, but is not practical for large-scale field studies; and serological assays are known to be rapid, cost-effective, and high throughput, but are less sensitive than RT-PCR (Gil et al., 2013). To overcome these issues, we prepared five highly specific and sensitive MAbs specific for PVS and developed three Olodaterol serological assays for PVS detection, which should be of use in large scale epidemiological studies and potato breeding. 2.?Materials and methods 2.1. Viruses and field potato samples Potato leafroll virus (PLRV)-infected, ordinary strain of PVS (PVSO)-infected, and virus-free potato seedlings were kindly supplied by the Institute of Biotechnology and Germplasm Resources, Yunnan Provincial Academy of Agricultural Sciences (Kunming, Olodaterol China). Potato virus A (PVA), Potato virus Y (PVY) and PVX were previously characterized and maintained in potato plants. Field potato samples showing virus-like symptoms were collected from Yunnan Province, China. Virus in the PVSO-infected potato plants was purified according to a previously described method (Zhou et al., 1994) and was used as the immunogen for anti-PVS MAb preparation. 2.2. MAb preparation Purified PVS virion was used to immunize five BALB/c female mice as described previously by Shang et al. (2011). Hybridoma cells secreting anti-PVS MAbs and ascitic fluids containing MAbs were prepared as described by Wu et al. (2013a). Crude extract from PVS-inoculated plants was used as the coating antigen during an indirect-enzyme-linked immunosorbent assay (ELISA) to determine the titers of the resulting ascitic fluids. The isotypes of MAbs were discriminated with a mouse MAb isotyping kit as described (Sigma-Aldrich, St. Louis, MO, USA). The specificity and sensitivity of the.