Expression profiling by small RNA-seq for identifying miRNA associated with sound vibration in plant

Plant can perceive and respond natural sound vibration (SV). Artificial SV also served as a novel trigger of induced resistance, although approaches for activating such plant innate immunity intensively studied on the use of biological and chemical agents (BCA). Artificial SV pre-treatment protected Arabidopsis thaliana seedlings against insect pests and fungal pathogens. However, SV-mediated epigenetic modulation remains unexplored while CBA-mediated induced resistance is known as a complicated process involving epigenetic regulation. Here, we performed an expression profiling basd on small RNA-seq experiment to understand the role of 10 kHz SV-mediated epigenetic modification in induced resistance against a soil-borne pathogenic bacterium Ralstonia solanacearum. A group of plants was prepared to be expose to sound vibration (SV, 10 kHz) at 90 dB every 3 h for 10 days. The other group of plants with non-treatment were also prepared as a control group. Then, all samples were drenched with a pathogen (R. solanacearum). miRNA quantification data were generated by performing small RNA-seq analysis of 18 arabidopsis samples, which were obtained at 0d, 1d and 2d after dreching pathogen. Total RNA was isolated using the mirVana miRNA isolation kit (Ambion). Small RNAs (20 - 30 nt) were purified from 15% Novex TBE-Urea gel (Invitrogen) and ligated first with the 5’ RNA adaptor and then with the 3’ RNA adaptor provided by Illumina TruSeq small RNA sample preparation protocol. In each step, the ligated product was PAGE-gel purified. After first-strand synthesis and 11 cycles of PCR amplification, the product was PAGE-gel purified. Sequencing was performed in single end reads (75 bp) using NextSeq 500 platform (Illumina).