Fig. 2

Key steps of the ALSV-based VIGS protocol in NLL and its application for the silencing of LaPDS. A Virion production via agroinfiltration in N. benthamiana. The image shows representative plants 10–15 days post-agroinfiltration with either an empty Agrobacterium strain (mock-infiltrated) or a mixture of strains carrying pALSV-RNA1u, pALSV-RNA2u with or without target gene fragment, and pEAQ-HT-DEST1 (for the expression of P19) (ALSV-infiltrated). Only the ALSV-infiltrated plant shows viral symptoms such as stunting and necrosis on leaves and petioles. i, infiltrated leaves; s, systemic leaves. B Rub-inoculation of young NLL plants with N. benthamiana leaf sap from ALSV-infiltrated plants. C Effective silencing of LaPDS in NLL. The image shows representative NLL plants inoculated with ALSV without a target gene fragment (ALSV-0) or with a fragment of LaPDS (ALSV-PDS). ALSV-PDS-infected plants typically develop bleaching about 5 weeks after inoculation. D Normalized relative expression of LaPDS in plants infected with ALSV carrying a fragment of GFP (ALSV-GFP; control) or ALSV-PDS. Box plots represent data for five biological replicates. Whiskers represent values within 1.5 times the interquartile range over or under the 75th and 25th percentiles, respectively. Dots represent the individual data points, the dashed line represents the mean of ALSV-GFP, and the asterisks indicate a significant difference (two-sided Wilcoxon test; P ≤ 0.01)