Immunodiagnosis of groundnut and watermelon bud necrosis viruses using polyclonal antiserum to recombinant nucleocapsid protein of Groundnut bud necrosis virus

Jain, R. K.; Pandey, Amar Nath; Krishnareddy, M.; Mandal, Bikash

doi:10.1016/j.jviromet.2005.06.018

Cited by 42 publications

(24 citation statements)

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“…The major expressed protein appeared in the insoluble fraction, which could be successfully purified. Similar results of expression of recombinant His-tagged CP in the insoluble fraction have been reported for several plant viruses, pelargonium zonate spot virus [27], faba bean necrotic yellows virus [15], sugarcane yellow leaf virus [13], potato mop-top virus [5,6] and groundnut bud necrosis virus [12]. However, for some plant viruses such as sugarcane streak mosaic virus, cucurbit yellow stunting disorder virus, cymbidium mosaic virus, the His-tagged viral CP was expressed in soluble fraction [10,11,16,23].…”

Section: Discussionsupporting

confidence: 79%

Molecular characterization of Indian isolate of peanut mottle virus and immunodiagnosis using bacterial expressed core capsid protein

et al. 2014

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Section: Discussionsupporting

confidence: 79%

Molecular characterization of Indian isolate of peanut mottle virus and immunodiagnosis using bacterial expressed core capsid protein

et al. 2014

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“…Maximum expression of the fusion protein from pMal-PY-CP construct was achieved in E. coli TB1 cells. Higher induction was observed with 0.4 mM IPTG at 37°C for 3 h. Unlike other vector systems [1,6], the fusion protein in the present study was found in the soluble fraction. Whereas the pET-PY-CP clone did not show any visible expression under above conditions.…”

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confidence: 64%

Section: Abstract Pumpkin Yellow Vein Mosaic Virusmentioning

confidence: 99%

“…To overcome these limitations, molecular biology techniques are currently being used to express the viral genes of interest in E. coli and the recombinant antigen is used for the production of PAb. Recombinant antigens for several RNA plant viruses were successfully produced in E. coli [6,7,10], whereas, very few DNA plant viruses have been used for successful expression of recombinant antigen in E. coli [1].Pumpkin yellow vein mosaic virus, a bipartite begomovirus is known in India since 1950s [13] and the virus was characterized based on host range and serology [8]. Further, phylogenetic analysis of DNA-A sequence revealed that PYVMV was synonymous to squash leaf curl China virus (SLCCNV) reported from China [11,12].…”

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confidence: 99%

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Fusion coat protein of pumpkin yellow vein mosaic virus with maltose binding protein: Applications in immunodiagnosis of begomoviruses

et al. 2014

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Availability of adequate quantity of purified virus preparation from plant tissue is the major limitation in producing polyclonal antibodies (PAb) to begomovirus. Very few examples show successful utilization of E. coli expressed recombinant coat protein (CP) for immuno diagnosis of begomoviruses. In the present study, *771 bp CP gene (*29.0 kDa) of Pumpkin yellow vein mosaic virus (PYVMV) was expressed as a *71.0 kDa fusion protein with maltose binding protein (MBP) (*42.0 kDa) in E. coli. The MBP-CP was obtained in soluble state. The PAb to the purified fusion protein successfully detected PYVMV and other bipartite and monopartite begomoviruses in the field samples at 1:250 dilution in enzyme linked immunosorbent assay. Our study for the first time showed that MBP-tag fusion CP was suitable to produce diagnostic antibody to begomoviruses. Keywords Pumpkin yellow vein mosaic virusBegomoviruses (family Geminiviridae) are emerging constrain in the production of numerous crops [14]. Both, immune-and nucleo-based techniques have been employed for the detection of begomoviruses [5]. Begomoviruses are phloem limiting, generally not sap transmissible and many of them do not have suitable propagative hosts. As a result, transmission and purification of begomoviruses are difficult and yield of purified virus is usually low. However, tedious method of virus propagation in large number of host plants through whitefly (Bemisia tabaci) inoculation and purification of virus from large batches of infected plant tissues were performed to produce polyclonal antibodies (PAb) against a few begomoviruses such as squash leaf curl virus (SLCV) [3], tomato yellow leaf curl virus (TYLCV) [9] and tomato leaf curl Bangalore virus (ToLCBaV) [4]. The traditional method of begomovirus antigen preparation from infected plant tissue generally results into contamination with host proteins and thus limits their use in serological diagnosis. Begomovirus purification by conventional method further limits the renewable production of PAb. As a result, routine detection and identification using serology has been less commonly used for begomoviruses compared to the other plant viruses. To overcome these limitations, molecular biology techniques are currently being used to express the viral genes of interest in E. coli and the recombinant antigen is used for the production of PAb. Recombinant antigens for several RNA plant viruses were successfully produced in E. coli [6,7,10], whereas, very few DNA plant viruses have been used for successful expression of recombinant antigen in E. coli [1].Pumpkin yellow vein mosaic virus, a bipartite begomovirus is known in India since 1950s [13] and the virus was characterized based on host range and serology [8]. Further, phylogenetic analysis of DNA-A sequence revealed that PYVMV was synonymous to squash leaf curl China virus (SLCCNV) reported from China [11,12]. There is an increasing incidence of PYVMV and many other begomoviruses in the Indian subcontinent [14], but PAb to begomoviruses for immunodia...

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“…The recombinant approach is advantageous as it does not encounter the problems associated with the mixture of closely related viruses, purification and maintenance of live virus culture. This strategy has been successfully used for a number of plant viruses from different genera however not for carmoviruses [4,9,13,16]. To our knowledge, this is the first report of development of immunodiagnosis based on antiserum against bacterial expressed recombinant CP of any carmoviruses in general and SYMMV in particular.…”

Section: Discussionmentioning

confidence: 99%

Diagnosis of a new variant of soybean yellow mottle mosaic virus with extended host-range in India

et al. 2015

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Soybean yellow mottle mosaic virus (SYMMV, genus Carmovirus) was previously known to occur in South Korea and USA causing bright yellow mosaic in soybean. In this study, SYMMV (Car-Mb14 isolate) was isolated from mungbean (Vigna radiata) exhibiting mild mottling and puckering symptoms in the experimental field at Indian Agricultural Research Institute, New Delhi during 2012. The virus isolate, Car-Mb14 induced veinal mottling, mild mottling, chlorotic blotching, local and systemic necrosis in soybean, mungbean, blackgram, French bean and guar bean, respectively. The symptomatology of the present isolate of SYMMV was different from the previously reported South Korean isolate, as the later did not induce symptoms in any of the above legumes other than soybean. The present isolate was phylogenetically distinct and shared 90-93 % sequence identity in coat protein (CP) of 52 SYMMV isolates reported from Korea and USA. In order to know the serological relationships, the CP gene of the present isolate was over expressed as a 39 kDa protein in E. coli and an antiserum of 1:16,000 titer against the recombinant CP was produced. Serological cross reactivity analysis revealed that SYMMV was serologically related to blackgram mottle virus but not to cowpea mottle virus, the other legume infecting carmoviruses. The antiserum was used to detect prevalence of SYMMV in legume crops by ELISA. Out of 145 field samples of legumes (mungbean, blackgram, French bean and soybean) collected from different places in India, SYMMV was detected only in 16 samples of mungbean and one sample of blackgram. The natural infection of SYMMV in mungbean and blackgram was further confirmed based on CP gene sequence. This study provides evidence of occurrence of a new variant of SYMMV with distinct symptom phenotype and extended host-range in India.

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Immunodiagnosis of groundnut and watermelon bud necrosis viruses using polyclonal antiserum to recombinant nucleocapsid protein of Groundnut bud necrosis virus

Cited by 42 publications

References 6 publications

Molecular characterization of Indian isolate of peanut mottle virus and immunodiagnosis using bacterial expressed core capsid protein

Molecular characterization of Indian isolate of peanut mottle virus and immunodiagnosis using bacterial expressed core capsid protein

Fusion coat protein of pumpkin yellow vein mosaic virus with maltose binding protein: Applications in immunodiagnosis of begomoviruses

Diagnosis of a new variant of soybean yellow mottle mosaic virus with extended host-range in India

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