scholarly journals Transfer and Expression of Cucumber Mosaic Virus Coat Protein Gene in the Genome of Cucumis sativus

1991 ◽  
Vol 116 (6) ◽  
pp. 1098-1102 ◽  
Author(s):  
Paula P. Chee ◽  
Jerry L. Slightom
Keyword(s):  
Coat Protein ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Cucumis Sativus ◽  
Blot Analysis ◽  
Binary Vector ◽  
Enzyme Linked Immunosorbent Assay ◽  
Embryogenic Calli ◽  
Mature Embryos ◽  
Npt Ii

Cotyledon explants of cucumber (Cucumis sativus L. cv. Poinsett 76) seedlings were cocultivated with disarmed Agrobacterium strain C58Z707 that contained the binary vector plasmid pGA482GG/cpCMV19. The T-DNA region of this binary vector contains plant-expressible genes for neomycin phosphotransferase II (NPT II), β -glucuronidase (GUS), and the coat protein of cucumber mosaic virus strain C (CMV-C). After infection, the cotyledons were placed on Murashige and Skoog medium containing 100 mg kanamycidliter. Putative transformed embryogenic calli were obtained, followed by the development of mature embryos and their germination to plants. All transformed RO cucumber plants appeared morphologically normal and tested positive for NPT IL Southern blot analysis of selected cucumber DNAs indicated that NPT II, GUS, and CMV-C coat protein genes were integrated into the genomes. Enzyme-linked immunosorbent assay and Western blot analysis indicated that the CMV-C coat protein is present in the protein extracts of progeny plants. These results show that the Agrobacterium-mediated gene transfer system and regeneration via somatic embryogenesis is an effective method for producing transgenic plants in Cucurbitaceae.

scholarly journals First Report of Cucumber mosaic virus Subgroup II Infecting Lycopersicon esculentum in India

Plant Disease ◽  
2006 ◽  
Vol 90 (11) ◽  
pp. 1457-1457 ◽  
Author(s):  
N. Sudhakar ◽  
D. Nagendra-Prasad ◽  
N. Mohan ◽  
K. Murugesan
Keyword(s):  
Coat Protein ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Tamil Nadu ◽  
Indian Subcontinent ◽  
Enzyme Linked Immunosorbent Assay ◽  
Infected Leaf ◽  
Polymorphism Analysis ◽  
First Report ◽  
Subgroup Ii

During a survey in January 2006 near Salem in Tamil Nadu (south India), Cucumber mosaic virus was observed infecting tomatoes with an incidence of more than 70%. Plants exhibiting severe mosaic, leaf puckering, and stunted growth were collected, and the virus was identified using diagnostic hosts, evaluation of physical properties of the virus, compound enzyme-linked immunosorbent assay (ELISA) (ELISA Lab, Washington State University, Prosser), reverse-transcription polymerase chain reaction (RT-PCR), and restriction fragment length polymorphism analysis (DSMZ, S. Winter, Germany). To determine the specific CMV subgroup, total RNA was extracted from 50 infected leaf samples using the RNeasy plant RNA isolation kit (Qiagen, Hilden, Germany) and tested for the presence of the complete CMV coat protein gene using specific primers as described by Rizos et al. (1). A fragment of the coat protein was amplified and subsequently digested with MspI to reveal a pattern of two fragments (336 and 538 bp), indicating CMV subgroup II. No evidence of mixed infection with CMV subgroup I was obtained when CMV isolates representing subgroups I (PV-0419) and II (PV-0420), available at the DSMZ Plant Virus Collection, were used as controls. Only CMV subgroup I has been found to predominantly infect tomato in the Indian subcontinent, although Verma et al. (2) identified CMV subgroup II infecting Pelargonium spp., an ornamental plant. To our knowledge, this is the first report of CMV subgroup II infecting tomato crops in India. References: (1) H. Rizos et al. J. Gen. Virol. 73:2099, 1992. (2) N. Verma et al. J. Biol. Sci. 31:47, 2006.

scholarly journals Tobacco Plants Transformed with a Modified Coat Protein of Tomato Mottle Begomovirus Show Resistance to Virus Infection

1999 ◽  
Vol 89 (8) ◽  
pp. 701-706 ◽  
Author(s):  
X. H. Sinisterra ◽  
J. E. Polston ◽  
A. M. Abouzid ◽  
E. Hiebert
Keyword(s):  
Coat Protein ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Protein Gene ◽  
Binary Vector ◽  
Enzyme Linked Immunosorbent Assay ◽  
Chain Reaction ◽  
Resistance Response ◽  
Tobacco Plants ◽  
Polymerase Chain

Tobacco plants (Nicotiana tabacum ‘Xanthi’) were transformed with a binary vector containing the coat protein gene of tomato mottle begomo-virus (ToMoV) modified by the deletion of 30 nucleotides in the 5′ end. The R1 generation was screened for resistance to ToMoV by inoculation with viruliferous whiteflies. Fifteen days after inoculation, symptom development was recorded weekly for up to 120 days using a visual scale, and ToMoV infection was confirmed by polymerase chain reaction and enzyme-linked immunosorbent assay. The response to high inoculation levels of ToMoV varied and ranged from susceptibility to immunity. The transgene transcript was detected by northern blot analysis; however, the transgene product could not be detected by protein blot analysis using antisera reactive with ToMoV coat protein. The lack of detection of the transgene product in resistant plants suggests that it is not involved in eliciting the resistance response and that resistance may be mediated by the transgene transcript.

Coat protein-mediated resistance as an approach for controlling an Egyptian isolate of Cucumber mosaic virus (subgroup I)

Biologia ◽  
2008 ◽  
Vol 63 (5) ◽  
Author(s):  
Ali El-Borollosy ◽  
Sabry Mahmoud ◽  
Abdel-Sabour Khaled
Keyword(s):  
Coat Protein ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Restriction Enzymes ◽  
Significant Loss ◽  
Enzyme Linked Immunosorbent Assay ◽  
35S Promoter ◽  
Rt Pcr ◽  
Dot Blot ◽  
Egyptian Isolate

AbstractCucumber mosaic virus (CMV, cucumovirus) is the most important virus infecting cucurbit crops in Egypt and worldwide causing significant loss in yield quality and quantity. The main target of the present work was to establish a simple controlling system for an Egyptian isolate of such virus (belonging to the subgroup I) via production of tobacco transgenic plants expressing viral coat protein (CP). Coat protein gene (cp) was isolated and amplified using immunocapture-reverse transcriptase-polymerase chain reaction (IC-RT-PCR) and primers with add-on restriction sites for SmaI and SacI enzymes. The genes were cloned in pBI121 vector plasmid between the CaMV 35S promoter and the nos terminator after removing the Gus gene by restriction enzymes digestion. The new construct was used for Agrobacterium tumefaciens transformation, which was then used for tobacco transformation. Evaluation of transformation success and CP expression degree were confirmed using indirect enzyme-linked immunosorbent assay (I-ELISA) and dot blot immuno-binding assay (DBIA). PCR and RT-PCR were used to study the integration of cp within genetic plant system and to what extent this gene was transcript. It was concluded that in spite of integration success some transformed plants can transcript the gene more than the others do. Plants resistance was tested by challenging with CMV under study and remarkable success was obtained in plants with higher gene transcription and translation degree.

scholarly journals Transferring Cucumber Mosaic Virus-White Leaf Strain Coat Protein Gene into Cucumis melo L. and Evaluating Transgenic Plants for Protection against Infections

1994 ◽  
Vol 119 (2) ◽  
pp. 345-355 ◽  
Author(s):  
Carol Gonsalves ◽  
Baodi Xue ◽  
Marcela Yepes ◽  
Marc Fuchs ◽  
Kaishu Ling ◽  
...  
Keyword(s):  
New York ◽  
Coat Protein ◽  
Transgenic Plants ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Microprojectile Bombardment ◽  
Leaf Tissue ◽  
White Leaf ◽  
Npt Ii ◽  
Transfer Procedures

A single regeneration procedure using cotyledon explants effectively regenerated five commercially grown muskmelon cultivars. This regeneration scheme was used to facilitate gene transfers using either Agrobacterium tumefaciens (using `Burpee Hybrid' and `Hales Best Jumbo') or microprojectile bombardment (using `Topmark') methods. In both cases, the transferred genes were from the T-DNA region of the binary vector plasmid pGA482GG/cp cucumber mosaic virus-white leaf strain (CMV-WL), which contains genes that encode neomycin phosphotransferase II (NPT II), β-glucuronidase (GUS), and the CMV-WL coat protein (CP). Explants treated with pGA482GG/cpCMV-WL regenerated shoots on Murashige and Skoog medium containing 4.4 μm 6-benzylaminopurine (BA), kanamycin (Km) at 150 mg·liter-1 and carbenicillin (Cb) at 500 mg·liter-1. Our comparison of A. tumefaciens- and microprojectile-mediated gene transfer procedures shows that both methods effectively produce nearly the same percentage of transgenic plants. R0 plants were first tested for GUS or NPT II expression, then the polymerase chain reaction (PCR) and other tests were used to verify the transfer of the NPT II, GUS, and CMV-WL CP genes. This analysis showed that plants transformed by A. tumefaciens contained all three genes, although co-transferring the genes into bombarded plants was not always successful. R1 plants were challenge inoculated with CMV-FNY, a destructive strain of CMV found in New York. Resistance levels varied according to the different transformed genotypes. Somaclonal variation was observed in a significant number of R0 transgenic plants. Flow cytometry analysis of leaf tissue revealed that a significant number of transgenic plants were tetraploid or mixoploid, whereas the commercial nontransformed cultivars were diploid. In a study of young, germinated cotyledons, however, a mixture of diploid, tetraploid, and octoploid cells were found at the shoot regeneration sites.

scholarly journals Epidemic of Potato virus Y and Cucumber mosaic virus in Henan Province Tobacco

Plant Disease ◽  
2001 ◽  
Vol 85 (4) ◽  
pp. 447-447 ◽  
Author(s):  
X. D. Li ◽  
Y. Q. Li ◽  
H. G. Wang
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Potato Virus ◽  
Potato Virus Y ◽  
Early Stage ◽  
Potato Virus X ◽  
Henan Province ◽  
Enzyme Linked Immunosorbent Assay ◽  
Resistant Varieties ◽  
Vein Necrosis

Flue-cured tobacco is an important crop in Henan Province, China. During the 2000 growing season, many tobacco plants showed various degrees of mottling, mosaic, vein clearing, or vein necrosis in most of the counties. Some plants even died at an early stage of growth. A survey was conducted in May-June in several tobacco-growing counties, and the incidence of symptomatic plants in individual fields ranged from 10 to 85%. The most widely planted tobacco varieties, NC89, K326, and K346, were highly susceptible. Symptomatic plants were collected from Jiaxian and Xiangcheng counties and samples were tested by enzyme-linked immunosorbent assay for Tobacco mosaic virus (TMV), Cucumber mosaic virus (CMV), Potato virus Y (PVY), and Potato virus X (PVX). Of 65 samples tested, 21 were positive for only PVY, 16 positive for only CMV, one each was positive for only TMV or PVX. Nineteen samples were doubly infected with various combinations of these viruses and six were infected with combinations of three viruses. The causal agent(s) in the remaining sample could not be determined. In total, CMV was detected in 40 samples, PVY in 38, PVX in 10, and TMV in 7 samples. TMV and CMV used to be the most important viruses and PVY occurred only rarely. But PVY has become prevalent in Henan and in neighboring Shandong province (2). CMV and TMV were reported to be the most prevalent viruses in Shanxi (1) and Fujian Provinces (3). Because resistant varieties are not available, and mixed infections are more common, the results presented here explain why huge damage is occurring in tobacco crops in recent years. Some varieties are partially resistant to TMV and CMV but the varieties commonly grown are highly susceptible to PVY. Therefore, breeding for resistance to viruses, especially to PVY, is urgent to control the occurrence of tobacco viral diseases. References: (1) J. L. Cheng et al. Acta Tabacaria Sin. 4:43, 1998. (2) J. B. Wang et al. Chinese Tobacco Sci. 1:26, 1998. (3) L. H. Xie et al. Acta Tabacaria Sin. 2:25, 1994.

scholarly journals Allamanda Mosaic Caused by Cucumber mosaic virus in Taiwan

Plant Disease ◽  
2005 ◽  
Vol 89 (5) ◽  
pp. 529-529 ◽  
Author(s):  
Y. K. Chen ◽  
C. C. Yang ◽  
H. T. Hsu
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Chenopodium Quinoa ◽  
Rabbit Antiserum ◽  
Nucleotide Sequence Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rt Pcr ◽  
Indicator Plants

Allamanda (Allamanda cathartica L., family Apocynaceae) is native to Brazil and is a popular perennial shrub or vine ornamental in Taiwan. Plants showing severe mosaic, rugosity, and leaf distortion symptoms on leaves are common in commercial nurseries and private gardens. Examination of crude sap prepared from symptomatic leaves using an electron microscope revealed the presence of spherical virus particles with a diameter of approximately 28 nm. The virus was mechanically transmitted to indicator plants and induced symptoms similar to those incited by Cucumber mosaic virus (CMV). The virus caused local lesions on inoculated leaves of Chenopodium quinoa and C. amaranticolor and systemic mosaic in Cucumis sativus, Lycopersicon esculentum, Nicotiana benthamiana, N. glutinosa, N. rustica, and N. tabacum. On N. tabacum, necrotic ringspots developed on inoculated leaves followed by systemic mosaic. Tests of leaf sap extracted from naturally infected allamanda and inoculated indicator plants using enzyme-linked immunosorbent assay were positive to rabbit antiserum prepared to CMV. Viral coat protein on transblots of sodium dodecyl sulfate-polyacrylamide gel electrophoresis reacted with CMV subgroup I specific monoclonal antibodies (2). With primers specific to the 3′-half of RNA 3 (1), amplicons of an expected size (1,115 bp) were obtained in reverse transcription-polymerase chain reaction (RT-PCR) using total RNA extracted from infected allamanda and N. benthamiana. The amplified fragment (EMBL Accession No. AJ871492) was cloned and sequenced. It encompasses the 3′ part of the intergenic region of RNA 3 (158 nt), CP ORF (657 nt), and 3′ NTR (300 nt) showing 91.8–98.9% and 71.4–72.8% identities to those of CMV in subgroups I and II, respectively. Results of MspI-digested restriction fragment length polymorphism patterns of the RT-PCR fragment and the nucleotide sequence analysis indicate that the CMV isolate from allamanda belongs to subgroup IB, which is predominant on the island. To our knowledge, CMV is the only reported virus that infects allamanda and was first detected in Brazil (3), and this is the first report of CMV infection in allamanda plants occurring in Taiwan. References: (1) Y. K. Chen et al. Arch. Virol. 146:1631, 2001. (2) H. T. Hsu et al. Phytopathology 90:615, 2000. (3) E. W. Kitajima. Acta. Hortic. 234:451, 1988.

scholarly journals Cucumber Mosaic Virus, Tobacco Streak Virus, and Cucumber Mosaic Virus Satellite RNA Associated with Mosaic and Ringspot Symptoms in Ajuga reptans in Ohio

Plant Disease ◽  
1997 ◽  
Vol 81 (10) ◽  
pp. 1214-1214 ◽  
Author(s):  
J. R. Fisher ◽  
S. T. Nameth
Keyword(s):  
United States ◽  
Molecular Weight ◽  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
The United States ◽  
Negative Control ◽  
Enzyme Linked Immunosorbent Assay ◽  
Low Molecular Weight ◽  
Streak Virus ◽  
Ajuga Reptans

Creeping bugleweed (Ajuga reptans L.) is a perennial ornamental commonly grown as a ground cover in temperate climates. Commercial samples of the A. reptans cultivars Royalty, var. Atropurpurea Bronze, Bronze Beauty, and Burgundy Glow showing mosaic and ringspot symptoms were tested for the presence of virus infection by direct antibody sandwich enzyme-linked immunosorbent assay (ELISA) and viral-associated double-stranded (ds) RNA analysis. Cucumber mosaic cucumovirus (CMV) was detected by ELISA and dsRNA analysis in symptomatic samples of all cultivars tested. ELISA values were considered positive if the absorbance values were twice the negative control. Negative control values were established with asymptomatic tissue of the cv. Bronze Beauty. Tobacco streak ilarvirus (TSV) was detected only by ELISA in symptomatic samples of all cultivars except Royalty. No dsRNA suggestive of TSV was detected. Alfalfa mosaic virus (AMV) was detected by ELISA and dsRNA analysis in symptomatic samples of all cultivars tested except Royalty and var. Atropurpurea Bronze. dsRNA analysis also indicated the presence of a low molecular weight, possible satellite (sat) RNA associated with all symptomatic and asymptomatic Royalty and var. Atropurpurea Bronze plants tested. Northern (RNA) blot analysis with a digoxigenin-labeled full-length clone of the (S) CARNA-5 (-) CMV satRNA (ATCC no. 45124) confirmed that the low molecular weight RNA associated with the Royalty and var. Atropurpurea Bronze cultivars was indeed CMV satRNA. Only AMV has been previously reported in A. reptans in the United States (1). This is the first report of CMV and its satRNA, as well as TSV, in A. reptans in the United States. Reference: (1) W. T. Schroeder and R. Provvidenti. Plant Dis. Rep. 56:285, 1972.

Dalmatian Toadflax (Linaria dalmatica): New Host for Cucumber Mosaic Virus

2007 ◽  
Vol 21 (1) ◽  
pp. 41-44 ◽  
Author(s):  
Courtney L. Pariera Dinkins ◽  
Sue K. Brumfield ◽  
Robert K. D. Peterson ◽  
William E. Grey ◽  
Sharlene E. Sing
Keyword(s):  
Cucumber Mosaic Virus ◽  
Mosaic Virus ◽  
Aphid Transmission ◽  
Negative Control ◽  
Enzyme Linked Immunosorbent Assay ◽  
New Host ◽  
Tobacco Plants ◽  
Linaria Dalmatica ◽  
Mechanical Methods ◽  
Dalmatian Toadflax

To date, there have been no reports of Dalmatian toadflax serving as a host for cucumber mosaic virus (CMV). Infestations of Dalmatian toadflax may serve as a reservoir of CMV, thereby facilitating aphid transmission of CMV to both agricultural crops and native plants. The goal of this study was to determine whether Dalmatian toadflax is a host for CMV. Dalmatian toadflax seedlings were randomly assigned to two treatments (18 replicates/treatment): no inoculation (control) and inoculation with CMV (Fast New York strain). The Dalmatian toadflax seedlings were inoculated by standard mechanical methods and tested for the presence of CMV using enzyme-linked immunosorbent assay (ELISA). Ten of the 18 CMV-inoculated toadflax plants tested positive for the virus; 6 of the 18 displayed systemic mosaic chlorosis and leaf curling. All control plants tested negative. Transmission electron microscopy obtained from CMV-positive plants confirmed the presence of CMV based on physical properties. To verify CMV infestation, tobacco plants were assigned to the following treatments (six replicates/treatment): no inoculation (control), CMV-negative (control) inoculation, and a CMV-positive inoculation. Plants were inoculated by standard methods. Five of the 6 tobacco plants treated with the CMV-positive inoculum tested positive for CMV using ELISA. All control plants tested negative for the virus.

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