Spray application of a cocktail of dsRNAs reduces infection of chilli leaf curl virus in Nicotiana benthamiana

Singh, Oinam Washington; Gupta, Dipinte; Joshi, B. C.; Roy, Anirban; Mukherjee, Sunil Kumar; Mandal, Bikash

doi:10.1007/s41348-021-00549-5

Cited by 9 publications

(8 citation statements)

References 36 publications

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“…RNAi method will have to overcome some limitations like the ability to withstand natural field conditions and heavy loads of viruses, rapidly evolving ChiLCV variants that escaped sequence-specific recognition in RNAi mechanism, and also uncertainties about the release of genetically modified (GM) crops. Recently, Singh et al ( 2022 ) a novel strategy of application of a cocktail of dsRNAs in Nicotiana benthamiana was attempted to prevent ChiLCV infection through RNAi. In this study, under in vivo conditions separate three dsRNA molecules were prepared using E. coli strain HT115 with the three suppressor genes, C2, V2, and C4 of ChiLCV.…”

Section: Advanced Methodsmentioning

confidence: 99%

An Overview of Chili Leaf Curl Disease: Molecular Mechanisms, Impact, Challenges, and Disease Management Strategies in Indian Subcontinent

et al. 2022

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Leaf curl disease in a chili plant is caused mainly by Chili leaf curl virus (ChiLCV) (Family: Geminiviridae, Genus: Begomovirus). ChiLCV shows a widespread occurrence in most of the chili (Capsicum spp.) growing regions. ChiLCV has a limited host range and infects tomatoes (Solanum lycopersicum), potatoes (S. tuberosum), and amaranth (Amaranthus tricolor). The virus genome is a monopartite circular single-stranded DNA molecule of 2.7 kb and associated with α and β-satellites of 1.3 and 1.4 kb, respectively. The virus genome is encapsulated in distinct twinned icosahedral particles of around 18–30 nm in size and transmitted by Bemisia tabaci (Family: Aleyrodidae, Order: Hemiptera). Recently, bipartite begomovirus has been found to be associated with leaf curl disease. The leaf curl disease has a widespread distribution in the major equatorial regions viz., Australia, Asia, Africa, Europe, and America. Besides the PCR, qPCR, and LAMP-based detection systems, recently, localized surface-plasmon-resonance (LPSR) based optical platform is used for ChiLCV detection in a 20–40 μl of sample volume using aluminum nanoparticles. Management of ChiLCV is more challenging due to the vector-borne nature of the virus, therefore integrated disease management strategies need to be followed to contain the spread and heavy crop loss. CRISPR/Cas-mediated virus resistance has gained importance in disease management of DNA and RNA viruses due to certain advantages over the conventional approaches. Therefore, CRISPR/Cas system-mediated resistance needs to be explored in chili against ChiLCV.

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Section: Advanced Methodsmentioning

confidence: 99%

An Overview of Chili Leaf Curl Disease: Molecular Mechanisms, Impact, Challenges, and Disease Management Strategies in Indian Subcontinent

et al. 2022

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“…Following the methodology described by (61) with minor modifications (62), single colonies of E. coli HT115 transformants carrying the hpTR-1+2_L4440 plasmid were cultured at 37°C with shaking for 16 hours in LB medium supplemented with tetracycline (12.5 µg/mL) and ampicillin (100 µg/mL). Each culture was then diluted 1:100 in a final volume of 0.5 L of LB medium supplemented with the same antibiotics and incubated at 37°C until reaching an OD600 of 0.5.…”

Section: In Vivo Production Of Hprna In E Coli Ht115mentioning

confidence: 99%

“…Topical dsRNA application also shows promise against monopartite geminiviruses such as Tomato yellow leaf curl virus (TYLCV) (56) and Chilli leaf curl virus (ChiLCV) (57).…”

Section: Introductionmentioning

confidence: 99%

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Hairpin-RNA Spray Confers Resistance to Mungbean Yellow Mosaic India Virus in Mungbean

Dhobale,

Sahoo

2024

Preprint

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The prevalence of Begomovirus diseases poses a significant threat to legume crops, necessitating the exploration of innovative control measures. This investigation explores the utilization of dsRNA molecules to initiate RNA interference (RNAi) targeting begomovirus, particularly focusing on Mungbean yellow mosaic India virus (MYMIV) and its potential threat to mungbean crops. Given the lack of genetic resistance in commercially available mungbean varieties, the study endeavors to employ RNAi as a strategic method for the effective control of MYMIV. The approach involves the preparation of vectors for the transient expression of three dsRNA targeting multiple overlapping ORFs of MYMIV DNA A through agroinoculation, and the selection of a highly efficient construct for dsRNA expression in bacteria, enabling topical application to mungbean plants in growth chamber experiments. Agroinoculation assays demonstrate effective resistance against MYMIV, as confirmed by reduced symptom severity, limited virus accumulation, and the presence of viral mRNAs. The stability of the prepared dsRNA against nucleases is confirmed, showcasing its ability to enter plant cells, move to non treated trifoliate leaves, and form siRNA when sprayed onto mungbean leaves, as validated by qRT-PCR and northern blotting. Varied combinations of the timing of dsRNA spray and virus infection reveal differential resistance against the virus. Notably, spraying two days before or on the same day as virus exposure emerges as the most suitable time to achieve optimal resistance against virus infection. In light of these findings, the topical application of dsRNAs stands out as a promising and effective strategy for MYMIV control in mungbean crops.

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“…RNAi method will have to overcome some limitations like the ability to withstand natural field conditions and heavy loads of viruses, rapidly evolving ChiLCV variants that escaped sequence-specific recognition in RNAi mechanism, and also uncertainties about the release of genetically modified (GM) crops. Recently, Singh et al (2022) a novel strategy of application of a cocktail of dsRNAs in Nicotiana benthamiana was attempted to prevent ChiLCV infection through RNAi. In this study, under in vivo conditions separate three dsRNA molecules were prepared using E. coli strain HT115 with the three suppressor genes, C2, V2, and C4 of ChiLCV.…”

Section: Pathogen Derived Resistance (Pdr)mentioning

confidence: 99%

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Spray application of a cocktail of dsRNAs reduces infection of chilli leaf curl virus in Nicotiana benthamiana

Cited by 9 publications

References 36 publications

An Overview of Chili Leaf Curl Disease: Molecular Mechanisms, Impact, Challenges, and Disease Management Strategies in Indian Subcontinent

An Overview of Chili Leaf Curl Disease: Molecular Mechanisms, Impact, Challenges, and Disease Management Strategies in Indian Subcontinent

Hairpin-RNA Spray Confers Resistance to Mungbean Yellow Mosaic India Virus in Mungbean

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