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. 2017 Aug 21;7:8341. doi: 10.1038/s41598-017-08823-z

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Effect of chloride channel protein CLC-b-like mRNA and the 40S ribosomal protein S3a-like mRNA on the plant’s morphology. (A) The tomato plants were subjected to a knock-down assay by a VIGS technique using the TRV1 vector in combination with either the TRV2 empty vector, or with its derivatives. After 18-days post infiltration plants exhibited phenotypic alterations similar to viroid infection. TRV-EV, TRV empty vector inoculated plants; TRV-PDS, plants inoculated with pTRV2:PDS; TRV-CLC-b-like, the plant infiltrated with pTRV2:CLC-b-like; and, TRV-RPS3a, the plant infiltrated with pTRV2:RPS3a. (B) The total nucleic acids extracted from these plants were PCR analyzed for the systemic presence of TRV2 derivatives. Lane M, 1 kb plus molecular marker, Lane 1, TRV-EV; Lane 2, pTRV2:PDS; Lane 3, TRV-CLC-b-like; Lane 4, TRV-RPS3a; Lane 5, PCR positive control for the TRV-EV vector; and, lane 6, PCR negative control. Total RNA extracted samples from the agroinfitrated plants were analyzed by gel blot assay for the knock-down/suppression of (C) Chloride channel CLC-b-like and, (D) RPS3a-like mRNAs using gene specific radiolabeled probes. The 5 S rRNA was used as a loading control. Full size gel blots are presented in Fig. S1. (E) The gel blot signals from (C) and (D) were quantified and expressed as a ratio of the target mRNA to the 5S rRNA signals. For each set of experiments, the ratio of target mRNA to 5S rRNA obtained with TRV-EV (control) was set at a value of 1. The additional bars indicate the relative target mRNA/5S rRNA ratio for each TRV-derivate vector (as indicated) expressed. Each experiment was performed at least three times. Error bars indicate SD.