Avocado Root Transcriptome Analysis

Avocado roots from control, mild-WS, and severe-WS plants were harvested at t₁ and t₂. Three biological replicates per timepoint, in which each biological replicate consisted of three plants (n = 9), were used for RNA extraction in each experimental group. RNA from ground root tissue was extracted using the CTAB extraction method [161 (link)] with modification described by Zumaquero et al. [21 (link)]. RNA parameters and integrity were checked using a NanoDrop® ND-1000 (Nanodrop Technologies Inc., Wilmington, DE, USA) spectrophotometer based on the A260/280 and A260/230 wavelength ratios and running samples on a 2% agarose gel. RNA samples were treated with a DNase treatment with 1 U of RNase-free DNase (Thermo Scientific, Life Technologies Inc., Carlsbad, CA, USA), 1 μL of 10× reaction buffer with MgCl₂, 1 μg of RNA, 0.5 μL of RiboLock RNase Inhibitor (Thermo Scientific Inc., CA, USA), and diethylpyrocarbonate-treated water to a final volume of 10 μL in all RNA samples. The mixture was incubated according to the manufacturer’s instructions at 37 °C for 45 min followed by the addition of 1 μL of 50 mM EDTA and incubation at 65 °C for 10 min according to the manufacturer’s instructions.