Deteksi Cepat Carnation mottle virus pada Tanaman Anyelir (Dianthus caryophyllus L.)

Erniawati Diningsih; Gede Suastika; Tri Asmira Damayanti; Slamet Susanto

doi:10.21082/jhort.v27n1.2017.p95-104

Deteksi Cepat Carnation mottle virus pada Tanaman Anyelir (Dianthus caryophyllus L.)

Jurnal Hortikultura ◽

10.21082/jhort.v27n1.2017.p95-104 ◽

2017 ◽

Vol 27 (1) ◽

pp. 95

Author(s):

Erniawati Diningsih ◽

Gede Suastika ◽

Tri Asmira Damayanti ◽

Slamet Susanto

Keyword(s):

Dianthus Caryophyllus ◽

Mottle Virus ◽

Rt Pcr ◽

One Step ◽

Carnation Mottle Virus

Carnation mottle virus (CarMV) merupakan virus penting pada tanaman anyelir di Indonesia maupun di dunia. Deteksi virus yang mudah dan cepat, diperlukan untuk memantau sumber induk anyelir bebas virus. Tujuan penelitian adalah mengevaluasi tiga metode preparasi RNA total yang mudah dan cepat dari tanaman anyelir sebagai templat one step RT-PCR. Sumber RNA total adalah dari daun dan batang anyelir terinfeksi CarMV. Metode yang dievaluasi yaitu SDT, SEM, dan kit komersial sebagai pembanding. Optimasi dilakukan terhadap konsentrasi akhir primer (0.4 – 1.0 µM) dan MgCl2 (1.5 dan 2.0 mM). Metode SDT dan SEM berhasil mendapat RNA total dari tanaman anyelir baik dari sampel daun maupun batang. Keberhasilan yang didapat dengan metode SDT dan SEM sebanding dengan kit komersial. One step RT-PCR RNA total yang digabungkan dengan metode SDT dan SEM menghasilkan intensitas DNA yang sebanding dengan kit komersial. RNA total dari daun sebagai sumber templat one step RT-PCR terbaik dibandingkan batang. Preparasi RNA total dengan metode SDT dan SEM adalah metode cepat, mudah, dan murah dalam menyediakan templat one step RT-PCR. Konsentrasi primer 0.4 µM dan MgCl2 2 mM merupakan konsentrasi optimum dan menghasilkan hasil amplifikasi terbaik

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Eliminasi Carnation Mottle Virus Menggunakan Senyawa Antiviral pada Kultur Jaringan Anyelir (Dianthus caryophyllus L.)

Jurnal Hortikultura ◽

10.21082/jhort.v25n3.2015.p229-237 ◽

2016 ◽

Vol 25 (3) ◽

pp. 229

Author(s):

Erniawati Diningsih ◽

Gede Suastika ◽

Tri Asmira Damayanti ◽

S Susanto

Keyword(s):

Dianthus Caryophyllus ◽

Mottle Virus ◽

Carnation Mottle Virus

Carnation mottle virus (CarMV) merupakan salah satu virus penting pada tanaman anyelir dan semua kultivar anyelir yang ditanam di Jawa Barat terinfeksi oleh virus ini. Penelitian bertujuan mendapatkan metode eliminasi CarMV yang efektif untuk membebaskan planlet anyelir dari virus. Inisiasi eksplant terinfeksi dilakukan pada media MSO dan perbanyakan planlet dilakukan pada media perbanyakan MS yang mengandung 1,0 ppm BA dan 0,5 ppm kinetin (MSZ). Metode eliminasi CarMV yang diuji terdiri atas perlakuan 2-thiourasil dan amantadin dengan konsentrasi masing-masing 0, 5, 10, 15, 20, 25, dan 30 ppm, dan ribavirin 5 ppm sebagai pembanding. Tunas apikal planlet ditanam pada media perlakuan (MSZA). Setelah tunas tumbuh, meristem terminal diambil 0,5 mm untuk ditanam pada media MSZ. Kultur meristem terminal dari planlet pada perlakuan 2- thiourasil menghasilkan planlet bebas virus sebesar 0 – 57%. Konsentrasi 2-thiourasil 25 ppm menghasilkan persentase planlet bebas virus tertinggi, namun perlakuan tersebut toksik pada tanaman. Perlakuan amantadin menghasilkan 25,0 – 54,55% planlet bebas virus. Di antara perlakuan yang diuji, perlakuan antiviral amantadin dengan konsentrasi 5 – 30 ppm lebih optimal menghasilkan planlet anyelir bebas CarMV dan tidak toksik terhadap tanaman. Perlakuan amantadin 5 – 20 ppm mampu menghambat virus lebih tinggi dibandingkan perlakuan 2-thiourasil pada konsentrasi yang sama. Amantadin 5 – 30 ppm menghasilkan tingkat penghambatan virus sebesar 42,94 – 59,57%, sedangkan 2-thiourasil sebesar -8,18 – 63,03%. Senyawa 2-thiourasil dan amantadin berpotensi sebagai agen antiviral untuk mendapatkan tanaman anyelir bebas CarMV.

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Carnation mottle virus. [Distribution map].

Distribution Maps of Plant Diseases ◽

10.1079/dmpd/20073215038 ◽

2007 ◽

Author(s):

Keyword(s):

Central America ◽

Dianthus Caryophyllus ◽

Uttar Pradesh ◽

Mottle Virus ◽

Distribution Map ◽

Calla Lily ◽

Virus Distribution ◽

New Distribution ◽

Distribution In Europe ◽

Carnation Mottle Virus

Abstract A new distribution map is provided for Carnation mottle virus. Tombusviridae: Carmovirus. Hosts: carnation (Dianthus caryophyllus), sweet william (D. barbatus), Begonia elatior [B. hiemalis], B. cheinamantha, Daphne odorata, calla lily (Zantedeschia sp.) and garden lettuce (Lactuca sativa). Information is given on the geographical distribution in Europe (Belgium, Bulgaria, Czech Republic, Denmark, Finland, France, Germany, Hungary, Italy, Latvia, Lithuania, Moldova, Netherlands, Poland, Romania, Russia, Serbia, Spain, UK and Ukraine) and Asia (Fujian, Hebei, Jiangsu, Yunnan and Zhejiang, China; Gujarat and Uttar Pradesh, India; Iran, Israel; Honshu, Japan; Korea Republic; Lebanon; and Taiwan), Africa (Egypt), North America (British Columbia and Ontario, Canada; Mexico; and California, Colorado, Ohio, Pennsylvania and Wisconsin, USA), Central America and Carribean (Cuba), South America (Argentina, Chile, Colombia, Venezuela, and São Paulo, Brazil) and Oceania (Victoria, Australia, and New Zealand).

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Kloning Gen Coat Protein (CP) Carnation Mottle Virus (CarMV) pada Vektor Ekspresi [Cloning of Carnation Mottle Virus (CarMV) Coat Protein Gene into Expression Vector]

Jurnal Hortikultura ◽

10.21082/jhort.v31n1.2021.p51-60 ◽

2021 ◽

Vol 31 (1) ◽

pp. 51

Author(s):

Erniawati Diningsih

Keyword(s):

Coat Protein ◽

Expression Vector ◽

Rna Extraction ◽

Mottle Virus ◽

Rt Pcr ◽

Bacterial Host ◽

Colony Pcr ◽

Functional Clone ◽

Suitable Expression ◽

Carnation Mottle Virus

Carnation mottle virus (CarMV) termasuk anggota genus Carmovirus dalam famili Tombusviridae. Virus ini banyak ditemukan menginfeksi tanaman anyelir di Jawa Barat dan menyebabkan gejala mottle. Sebagai langkah awal untuk memproduksi antiserum melalui teknik ekspresi gen CP perlu diklon pada vektor yang sesuai. Penelitian ini bertujuan mendapatkan klon CarMV yang berfungsi melalui kloning dan subkloning gen CP CarMV ke dalam vektor ekspresi yang sesuai. Penelitian dilakukan dalam beberapa tahap, yaitu ekstraksi RNA total dan amplifikasi cDNA CarMV dengan RT-PCR, menggunakan primer spesifik CarMVF dan CarMVR yang mengandung situs enzim restriksi XhoI dan BamHI, kloning dan subkloning DNA sisipan, serta konfirmasi transforman. Rekombinan gen sisipan CP CarMV dalam bakteri dikonfirmasi dengan koloni PCR. Gen CP CarMV berhasil dikloning ke dalam TA vektor pTZ57R/T dan disubkloning ke vektor ekspresi pET28a. Sekuen rekombinan CP CarMV berhasil dikonfirmasi melalui perunutan DNA. Penelitian lebih lanjut diperlukan untuk mendapatkan produksi antigen rekombinan yang melimpah pada bakteri ekspresi dan kondisi yang sesuai.KeywordsDianthus caryophillus L.; Carmovirus; Kloning; Subkloning; Bakteri ekspresiAbstractCarnation mottle virus (CarMV) is a type member of Carmovirus genus in family of Tombusvirus. The virus infects carnation plants in the centre area production of West Java and it cause mottle symptoms. The research aimed to obtain functional clone(s) of CarMV CP gene in suitable expression kloning vector. The research was carried out through several steps, namely total RNA extraction and amplification of cDNA of CP CarMV by RT-PCR using specific primer pairs CarMVF and CarMVR containing restriction enzyme sites XhoI and BamHI, respectively, TA cloning, and subcloning into expression vector pET28a and confirmation of recombinant plasmids by colony PCR. CarMV CP gen was successfully cloned into TA cloning vector pTZ57R/T and subcloned into vector pET28a, alsowere confirmed by DNA sequencing. Future experiment is necessary to be conducted to obtain abundance recombinant antigen production of CarMV CP in suitable expression condition and bacterial host.

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EFFECTS OF ELECTRIC CURRENT VALUE AND TIME TREATMENTS ON ELIMINATION OF CARNATION MOTTLE VIRUS AND IN VITRO PLANTLET REGENERATION OF CARNATION (DIANTHUS CARYOPHYLLUS)

Acta Horticulturae ◽

10.17660/actahortic.2009.829.63 ◽

2009 ◽

pp. 395-398

Author(s):

S. Sepahpour ◽

A. Moieni ◽

M. Shamsbakhsh ◽

A. Baghizadeh

Keyword(s):

Electric Current ◽

Dianthus Caryophyllus ◽

Plantlet Regeneration ◽

Mottle Virus ◽

Current Value ◽

Carnation Mottle Virus

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First Report of Carnation mottle virus in Calla lily (Zantedeschia spp.)

Plant Disease ◽

10.1094/pdis.2003.87.12.1539c ◽

2003 ◽

Vol 87 (12) ◽

pp. 1539-1539 ◽

Cited By ~ 3

Author(s):

C.-C. Chen ◽

W.-F. Ko ◽

C.-Y. Lin ◽

F.-J. Jan ◽

H. T. Hsu

Keyword(s):

Mosaic Virus ◽

Polyacrylamide Gel Electrophoresis ◽

Dianthus Caryophyllus ◽

Spherical Particles ◽

Mottle Virus ◽

Enzyme Linked Immunosorbent Assay ◽

Infected Leaf ◽

First Report ◽

Cp Gene ◽

Carnation Mottle Virus

Calla lilies are ornamental plants of major economic importance in Taiwan. They are grown in the central and northern areas of the island, and ≈3 million stems are shipped annually. Calla lilies are susceptible to several viruses (1). Infections by Cucumber mosaic virus, Dasheen mosaic virus, Turnip mosaic virus, and Watermelon silver mottle virus were reported in Taiwan. Recently, virus-like symptoms including yellow mottling, light yellow spot, yellow ringspot, and mosaic were observed on leaves of field-grown calla lilies from Changhua County, located in central Taiwan. In March 2001, a virus culture was isolated from diseased calla lilies and established in Chenopodium quinoa Willd. and Nicotiana benthamiana Domin. When inoculated with the virus, healthy calla lilies developed chlorotic spots that enlarged and fused to form large, yellow patches on inoculated leaves. Symptoms were similar to those on the naturally infected plants observed in the fields. The virus induced chlorotic local lesions on C. quinoa, C. ficifolium Sm., C. amaranticolor Coste & Reyn, Cucurbita moschata Duchesne ex Poir, Lisianthus russellianum (Don.) Griseb, Phaseolus angularis Wight, Vigna angularis Willd., and V. radiata (L.) Wilczek. In addition to the localized chlorotic spots on inoculated leaves, systemic invasion of the virus was also observed 8 to 10 days postinoculation in Dianthus caryophyllus L., D. chinensis L., and Glycine max Merr. In N. benthamiana, the only symptom observed was systemic wilting. Examination of 2% of uranyl-acetate-stained samples using electron microscopy revealed the presence of spherical particles ≈34 to 35 nm in diameter in crude extracts of leaves of diseased calla lilies, or infected C. quinoa. Similar particles were also observed in the cytoplasm but not in the nuclei in ultrathin sections of virus-infected leaf tissues of C. quinoa and N. benthamiana. Differential centrifugation followed by sucrose density gradient centrifugation of tissue extracts of infected C. quinoa yielded virions with similar size. Sodium dodesyl sulfate polyacrylamide gel electrophoresis of the purified virus showed a single structural polypeptide ith a Mr of 41.6 kDa. The viral antigen reacted positively with its homologous antiserum and an antiserum against Carnation mottle virus (CarMV; Agdia, Inc., Elkhart, IN) in double antibody sandwich enzyme-linked immunosorbent assay. Using primers 5′-CTCCATGGTCATGGAA(A/G)ATAAA GGAGAA and 3′-CAACAAATATCCTACACTGTCCTAGGTG specific to the coat protein (CP) gene of CarMV, an expected viral CP gene product of 1.05 kb was amplified by reverse transcription-polymerase chain reaction from total RNA isolated from infected N. benthamiana. Comparisons of the 1,047-nucleotide CP gene with those of 15 CarMV isolates available in GenBank showed 94.6 to 98.2% nucleotide identity and 94.8 to 96.8% amino acid identity. Results from current studies indicate that the virus infecting calla lilies is an isolate of CarMV. To our knowledge, this is the first report of CarMV infection in calla lilies. The occurrence of CarMV in calla lilies has direct implication for the economically important nursery and floral industry in Taiwan. Reference: (1) F. W. Zettler and R. D. Hartman. Dieffenbachia, Caladium, and Zantedeschia. Pages 464–470 in: Virus and Virus-Like Diseases of Bulb and Flower Crops. G. Loebenstein, R. H. Lawson, and A. A. Brunt, eds. John Wiley and Sons, West Sussex, U.K., 1995.

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Duplex-RT-PCR assay for the simultaneous detection and discrimination of Brome mosaic virus and Cocksfoot mottle virus in cereal plants

Molecular Biology Reports ◽

10.1007/s11033-021-06485-9 ◽

2021 ◽

Author(s):

Katarzyna Trzmiel

Keyword(s):

Mosaic Virus ◽

Simultaneous Detection ◽

Brome Mosaic Virus ◽

Mottle Virus ◽

Grass Species ◽

Replicase Gene ◽

Rt Pcr ◽

Pcr Products ◽

Cereal Plants ◽

Cocksfoot Mottle Virus

AbstractBrome mosaic virus (BMV) and cocksfoot mottle virus (CfMV) are pathogens of grass species including all economically important cereals. Both viruses have been identified in Poland therefore they create a potential risk to cereal crops. In this study, a duplex—reverse transcription—polymerase chain reaction (duplex-RT-PCR) was developed and optimized for simultaneous detection and differentiation of BMV and CfMV as well as for confirmation of their co-infection. Selected primers CfMVdiag-F/CfMVdiag-R and BMV2-F/BMV2-R amplified 390 bp and 798 bp RT-PCR products within coat protein (CP) region of CfMV and replicase gene of BMV, respectively. Duplex-RT-PCR was successfully applied for the detection of CfMV-P1 and different Polish BMV isolates. Moreover, one sample was found to be co-infected with BMV-ML1 and CfMV-ML1 isolates. The specificity of generated RT-PCR products was verified by sequencing. Duplex-RT-PCR, like conventional RT-PCR, was able to detect two viruses occurring in plant tissues in very low concentration (as low as 4.5 pg/µL of total RNA). In contrast to existing methods, newly developed technique offers a significant time and cost-saving advantage. In conclusion, duplex-RT-PCR is a useful tool which can be implemented by phytosanitary services to rapid detection and differentiation of BMV and CfMV.

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