GSM5966663
Root tissues were  dissected after 6h. All sampling were performed in triplicate. For each sample, about 1500 segments of the lower two-thirds of the seedling roots were pooled, flash frozen in liquid nitrogen and later homogenised with a polysome-extraction buffer (Thellmann et al, 2020). The suspension was centrifuged for 15 min (16 000 g, 4°C). An aliquot of the homogenate was used for Bulk-RNA-extraction by use of TRIzol reagent (Invitrogen). The remaining supernatant was incubated with GFP-Trap® Magnetic Agarose beads (Chromotek, Munich, Germany). Ribosomal bound RNA was obtained by immunoprecipitation with magnetic anti-GFP beads in accordance with the manufacturer's instructions and subsequently purified by use of TRIzol reagent (Invitrogen). Illumina NextSeq libraries were prepared from 2 µg of total RNA (bulk) or 100ng (Ribosome bound) after depleting the rRNA via Ribo-Zero Plus rRNA Depletion Kit (Illumina) and sequencing performed on NextSeq 500 flow cells (12 samples per cell).