Purification and serological analyses of tospoviral nucleocapsid proteins expressed by Zucchini yellow mosaic virus vector in squash

Chen, Tsung-Chi; Hsu, H. T.; Jain, R. K.; Huang, Ching-Wen; Lin, Chia-Chun; Liu, Fanglin; Yeh, Shyi-Dong

doi:10.1016/j.jviromet.2005.05.014

Cited by 26 publications

(23 citation statements)

References 57 publications

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“…NP is the most abundant structural protein of tospoviruses, and as such it has served well as a candidate for generation of antibodies. Indeed the majority of tospoviral monoclonal and polyclonal antibodies are directed against this structural protein of the virus (Chen et al 2005;Jain et al 2005;Wu et al 2009). Current classification of tospoviruses based on serological relationship of NPs, has divided them into three related serogroups including Iris yellow spot virus (IYSV), Tomato spotted wilt virus (TSWV) and Watermelon silver mottle virus (WSMoV) (Chen et al 2010).…”

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confidence: 99%

Generation of polyclonal antibodies and serological analyses of nucleocapsid protein of Soybean vein necrosis-associated virus: A distinct soybean infecting tospovirus serotype

et al. 2012

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Soybean vein necrosis-associated virus (SVNaV) is a newly isolated tospovirus from fieldgrown soybeans in the United States. Polyclonal antibodies generated against the recombinant Escherichia coli-expressed nucleocapsid protein (NP) of the virus, reacted specifically with SVNaV and exhibited low, if any, cross-reactivity with other species within the genus Tospovirus. The serological results are in agreement with low sequence homology of the SVNaV-NP gene when compared with those of other tospovirus species, some of which are capable of infecting soybean naturally. Phylogenetic analysis utilizing NP sequences from several SVNaV isolates collected from different geographical regions and various soybean genotypes over 4 years showed close relationships, but distinct from the representatives of all the established serogroups or distinct serotypes within the genus of Tospovirus. All SVNaV isolates examined reacted strongly with the generated polyclonal antibodies. Collectively, our serological analyses suggest that SVNaV represents a new and distinct soybean-infecting tospovirus serotype. Furthermore, our data demonstrate that antiserum against NP has the potential to serve as a reliable probe for SVNaV detection in field-grown soybeans.

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confidence: 99%

Generation of polyclonal antibodies and serological analyses of nucleocapsid protein of Soybean vein necrosis-associated virus: A distinct soybean infecting tospovirus serotype

et al. 2012

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“…NP is an important functional protein and a major criterion for the classification of tospoviruses (Goldbach and Kuo, 1996;Chen et al, 2005). Antisera against NPs are usually utilized for diagnosis and identification of tospoviruses (Yeh et al, 1996;Ghotbi et al, 2005;Lin et al, 2005;Chen et al, 2010).…”

Section: Discussionmentioning

confidence: 99%

“…Several methods have been reported to prepare tospoviral NPs as immunogens, such as purification of tospoviruses or nucleocapsids directly from virus-infected plant tissues, expression of NP using a plant viral vector in planta or use of prokaryotic expression systems (Yeh et al, 1996;Chu et al, 2001;Chen et al, 2005;Jain et al, 2005;Lin et al, 2005;Wu et al, 2009;Khatabi et al, 2012). Among these, expression of fusion proteins in E. coli is the most advantageous, as it lends itself to easy tospovirus protein purification and ease of manipulation, and the background values caused by contamination of plant proteins can be neglected (Elliott et al, 1996).…”

Section: Discussionmentioning

confidence: 99%

Development of polyclonal antibodies against nucleocapsid protein of watermelon silver mottle virus and their application to diagnostic

Wu¹,

W²,

Y³

et al. 2014

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Summary. -Watermelon silver mottle virus (WSMoV) is an emerging disease of cucurbit crops in South China. Production of high-quality antibodies is necessary for the development of serological methods for detection of this virus. The nucleocapsid protein (NP) gene of WSMoV was amplified from WSMoV-infected watermelon leaves by RT-PCR and cloned into vector pET-28a (+) for prokaryotic expression. After identification via enzyme digestion and sequencing, the recombinant clone was transformed into Escherichia coli Rosetta (DE3) for protein expression. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results showed that the molecular weight of the WSMoV NP fusion protein was 34.1 kDa. The fusion protein was purified and used as antigen for the preparation of polyclonal antisera in rabbits. Results of indirect ELISA and western blot analysis showed that the antisera reacted specifically with WSMoV NP. In addition, sensitivity and specificity of the antisera were examined on a number of infected field samples by indirect ELISA. These findings will facilitate further immunological and serological studies of WSMoV.Keywords: watermelon silver mottle virus; polyclonal antibody; western blot; prokaryotic expression Acta virologica 58: 167 -172, 2014 doi:10.4149/av_2014_02_167 * Corresponding author. E-mail: raoxq@hotmail.com; fax: 086-020-85281107. Abbreviations: CaCV = capsicum chlorosis virus; CMV = cucumber mosaic virus; HCRV = hippeastrum chlorotic ringspot virus; IPTG = Isopropyl-β-D-thiogalactopyranoside; NP = nucleocapsid protein; sodium WSMoV = watermelon silver mottle virus

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“…The sixteen viruses [Tomato spotted wilt virus, Tomato chlorotic spot virus, Groundnut ringspot virus (GRSV), Impatiens necrotic spot virus (INSV), Groundnut bud necrosis virus (GBNV), Watermelon silver mottle virus (WSMoV), Peanut yellow spot virus (PYSV), Zucchini lethal chlorosis virus, Chrysanthemum stem necrosis virus, Iris yellow spot virus, Peanut chlorotic fan-spot virus (PCFV), Melon yellow spot virus, Watermelon bud necrosis virus (WBNV), Tomato yellow fruit ring virus, Calla lily chlorotic spot virus (CCSV) and Capsicum chlorosis virus (CaCV)] included in the Tospovirus genus as definite or tentative species have been classified into three major serogroups (Tomato spotted wilt virus, WSMoV and Iris yellow spot virus) and four distinct serotypes (INSV, PYSV, PCFV and Melon yellow spot virus) based on the serological relationships and phylogenetical analysis of N protein (Chen et al, 2005a). Capsicum chlorosis virus was reported infecting capsicum and tomato in Australia and Thailand (Lee et al, 2002;Permachandra et al, 2005), and belongs to WSMoV serogroup (Chen et al, 2005b). The sequence of the N gene of an unpublished peanut strain (accession number AM087456) from Thailand shared 92.3% amino acid (aa) identity with that of CaCV Thailand tomato strain (CaCV-TT).…”

Section: Introductionmentioning

confidence: 99%

“…S RNA encodes a non-structural (NSs) protein in the viral (v) sense, and a nucleocapsid (N) protein in the viral complementary (vc) sense (de Haan et al, 1990 (Chen et al, 2005a). Capsicum chlorosis virus was reported infecting capsicum and tomato in Australia and Thailand (Lee et al, 2002;Permachandra et al, 2005), and belongs to WSMoV serogroup (Chen et al, 2005b). The sequence of the N gene of an unpublished peanut strain (accession number AM087456) from Thailand shared 92.3% amino acid (aa) identity with that of CaCV Thailand tomato strain (CaCV-TT).…”

Section: Introductionmentioning

confidence: 99%

Characterization of a New Strain of Capsicum chlorosis virus from Peanut (Arachis hypogaea L.) in China

Chen¹,

Xu²,

Yan³

et al. 2007

Journal of Phytopathology

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A new strain of Capsicum chlorosis virus (CaCV) from peanut (Arachis hypogaea L.) in China, designated CaCV-CP, was characterized. CaCV-CP causes yellow spots and necrosis on the leaves of affected peanut plants. Of 31 plant species inoculated mechanically, 24 were susceptible to this strain. Quasi-spherical virions were present in ultrathin section of diseased leaves. The complete sequence of S RNA of CaCV-CP consisted of 3399 nucleotides (nts). The NSs and N genes of the virus contained 1320 nts and 828 nts, respectively; these two open reading frames were in an ambisense arrangement. The N gene of CaCV-CP shared 84.7-86.4% and 92.4-93.1% identity with that of CaCV strains from Thailand and Australia at nt and amino acid levels, respectively (the GenBank accession number of the sequence reported in this study is DQ355974).

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Purification and serological analyses of tospoviral nucleocapsid proteins expressed by Zucchini yellow mosaic virus vector in squash

Cited by 26 publications

References 57 publications

Generation of polyclonal antibodies and serological analyses of nucleocapsid protein of Soybean vein necrosis-associated virus: A distinct soybean infecting tospovirus serotype

Generation of polyclonal antibodies and serological analyses of nucleocapsid protein of Soybean vein necrosis-associated virus: A distinct soybean infecting tospovirus serotype

Development of polyclonal antibodies against nucleocapsid protein of watermelon silver mottle virus and their application to diagnostic

Characterization of a New Strain of Capsicum chlorosis virus from Peanut (Arachis hypogaea L.) in China

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