Abstract
Downregulation of AM fungal genes using a plant viral vector is feasible. A partial sequence of a target fungal gene is cloned into the multicloning site of CMV2-A1 vector developed from RNA2 of Cucumber mosaic virus Y strain, and the RNA2, together with RNA1 and RNA3 of the virus, are in vitro-transcribed. Inoculation of Nicotiana benthamiana with these viral RNAs results in reconstitution of the virus in the plant, which triggers silencing of the fungal gene. Here, we describe not only the methods but also several tips for successful application of virus-induced gene silencing to AM fungi.
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References
Gerdemann JW (1968) Vesicular-arbuscular mycorrhiza and plant growth. Annu Rev Phytopathol 6:397–418
Nowara D, Gay A, Lacomme C et al (2010) HIGS: host-induced gene silencing in the obligate biotrophic fungal pathogen Blumeria graminis. Plant Cell 22:3130–3141
Wang M, Weiberg A, Lin F-M et al (2016) Bidirectional cross-kingdom RNAi and fungal uptake of external RNAs confer plant protection. Nat Plants 2:16151
Andika IB, Wei S, Cao C et al (2017) Phytopathogenic fungus hosts a plant virus: a naturally occurring cross-kingdom viral infection. Proc Natl Acad Sci U S A 114:12267–12272
Helber N, Wippel K, Sauer N et al (2011) A versatile monosaccharide transporter that operates in the arbuscular mycorrhizal fungus Glomus sp. is crucial for the symbiotic relationship with plants. Plant Cell 23:3812–3823
Sahu PP, Puranik S, Khan M, Prasad M (2012) Recent advances in tomato functional genomics: utilization of VIGS. Protoplasma 249:1017–1027
Kikuchi Y, Hijikata N, Ohtomo R et al (2016) Aquaporin-mediated long-distance polyphosphate translocation directed towards the host in arbuscular mycorrhizal symbiosis: application of virus-induced gene silencing. New Phytol 211:1202–1208
Kanazawa A, J-I I, Shimura H et al (2011) Virus-mediated efficient induction of epigenetic modifications of endogenous genes with phenotypic changes in plants. Plant J 65:156–168
Nagamatsu A, Masuta C, Senda M et al (2007) Functional analysis of soybean genes involved in flavonoid biosynthesis by virus-induced gene silencing. Plant Biotechnol J 5:778–790
Otagaki S, Arai M, Takahashi A et al (2006) Rapid induction of transcriptional and post-transcriptional gene silencing using a novel Cucumber mosaic virus vector. Plant Biotechnol 23:259–265
Tasaki K, Terada H, Masuta C, Yamagishi M (2016) Virus-induced gene silencing (VIGS) in Lilium leichtlinii using the Cucumber mosaic virus vector. Plant Biotechnol 33:373–381
Palukaitis P, Garcia-Arenal F (eds) (2019) Cucumber mosaic virus. APS Press, St Paul, MN, p 380
Smith SE, Read DJ (2008) Mycorrhizal Symbiosis, 3rd edn. Academic Press, San Diego, CA, p 787
Suzuki M, Kuwata S, Kataoka J et al (1991) Functional analysis of deletion mutants of cucumber mosaic virus RNA3 using an in vitro transcription system. Virology 183:106–113
Varkonyi-Gasic E, Wu R, Wood M et al (2007) Protocol: a highly sensitive RT-PCR method for detection and quantification of microRNAs. Plant Methods 3:1–12
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Ezawa, T., Maruyama, H., Kikuchi, Y., Yokoyama, K., Masuta, C. (2020). Application of Virus-Induced Gene Silencing to Arbuscular Mycorrhizal Fungi. In: Ferrol, N., Lanfranco, L. (eds) Arbuscular Mycorrhizal Fungi. Methods in Molecular Biology, vol 2146. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0603-2_19
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DOI: https://doi.org/10.1007/978-1-0716-0603-2_19
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