GSM2731772: No Bacteria_minusP_30uM_exp1_rep3 [SC003]; Arabidopsis thaliana; RNA-Seq
Total RNA was extracted from Arabidopsis seedlings according to Logemann, et al. (PMID: 2441623). Frozen seedlings were pulverized in liquid nitrogen. Samples were homogenized in 400 µl of Z6-buffer; 8 M guanidinium-HCl, 20 mM MES, 20 mM EDTA pH 7.0. Following the addition of 400 µl phenol:chloroform:isoamylalcohol; 25:24:1, samples were vortexed and centrifuged (20000 g, 10 min) for phase separation. The aqueous phase was transferred to a new 1.5 ml tube and 0.05 volumes of 1N acetic acid and 0.7 volumes 96 % ethanol were added. The RNA was precipitated at -20 °C overnight. Following centrifugation, (20,000 g, 10 min, 4 °C) the pellet was washed with 200 µl sodium-acetate (pH 5.2) and 70 % ethanol. The RNA was dried, and dissolved in 30 µl of ultrapure water and stored at -80 °C until use. poly(A) RNA was isolated from 1000 ng total RNA and used to prepare non-stranded Illumina RNA-seq libraries. Following library preparation, quality control and quantification were performed using a 2100 Bioanalyzer instrument (Agilent) and the Quant-iT PicoGreen dsDNA Reagent (Invitrogen), respectively.
GEO Accession
GSM2731772