Abstract
Solanum torvum has broad resistance to wilt-causing bacteria and fungi and nematodes. Here, we applied virus-induced gene silencing (VIGS) in S. torvum and optimized the experimental conditions to suppress a carotenoid biosynthesis gene with high efficiency. Using this optimized method, we suppressed the StPsbO and StPsbP genes, which encode components of the oxygen-evolving complex, in S. torvum. Silencing of StPsbO and StPsbP impaired defense response in S. torvum, making it more susceptible to Ralstonia solanacearum. These results indicate that the optimized VIGS method can be used to identify genes required for plant immunity in S. torvum.
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Funding
This work was supported by the Japan Society for the Promotion of Science[(to M.N. (JP18J02213 and JP19K15847)]; Yamazaki Spice Promotion Foundation (to M.N.); Fuji Foundation for Protein Research (to M.N.); and Institute for Fermentation, Osaka (to M.N.).
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10327_2021_1040_MOESM1_ESM.pdf
Figure S1: Silencing of PsbO and PsbP in Solanaceae plants. (a) Expression levels of NbPsbO and NbPsbP in leaves of Nicotiana benthamiana plants after tobacco rattle virus (TRV)-based virus-induced gene silencing (VIGS). Gene expression was analyzed using semi-quantitative RT-PCR. NbActin was used as the endogenous control. (b) Phenotypes of the control, NbPsbO-, and NbPsbP-silenced plants 3 weeks after VIGS. (c) Growth of VIGS plants. Stem length of plants in (b). Values are means ±SD of 10 replicates. Asterisks denote significant differences compared with the TRV:GUS control (P < 0.01, Dunnett’s test). (d) Chlorophyll content of the VIGS plants. Foliar chlorophyll content was measured for plants assessed in (b) using a chlorophyll meter. Values are means ± SD of 10 replicates. Asterisks denote statistically significant differences compared with the TRV:GUS control (P < 0.01, Dunnett’s test). (e) Expression levels of CaPsbO and CaPsbP in leaves of the VIGS Capsicum annuum plants. Gene expressions were analyzed using semi-quantitative RT-PCR. CaActin was used as the endogenous control. (f) Phenotypes of the control, CaPsbO-, and CaPsbP-silenced plants 5 weeks after VIGS. (g) Growth of the VIGS plants. Stem length was measured for plants in (f). Values are means ± SD of 10 replicates. Asterisks denote significant differences compared with the TRV:GUS control (P < 0.01, Dunnett’s test). (h) Chlorophyll content of VIGS plants. Foliar chlorophyll content was measured for the plants in (f) using a chlorophyll meter. Values are means ± SD of 10 replicates. Asterisks denote significant differences compared with the TRV:GUS control (P < 0.01, Dunnett’s test)
10327_2021_1040_MOESM2_ESM.pdf
Figure S2: Effect of PsbO- and PsbP-silencing on disease development in Solanaceae plants. (a) Daily disease index for wilt on the control, NbPsbO-, and NbPsbP-silenced Nicotiana benthamiana plants after inoculation with Ralstonia solanacearum. (b) Photographs of plants assessed in (a) at 9 days after inoculation. (c) Daily disease index for wilt on the control, CaPsbO-, and CaPsbP-silenced Capsicum annuum plants after inoculation with R. solanacearum. Stems of silenced plants were inoculated with R. solanacearum, and wilt was scored daily. (d) Photographs of plants assessed in (c) at 6 days after inoculation. All experiments were done three times with similar results; representative results are shown
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Nakano, M., Mukaihara, T. Virus-induced gene silencing in Solanum torvum. J Gen Plant Pathol 88, 10–16 (2022). https://doi.org/10.1007/s10327-021-01040-7
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DOI: https://doi.org/10.1007/s10327-021-01040-7