sRNA-seq of Arabidopsis thaliana Col-0 (nrpd1-/-) seeedlings
Total RNA was extracted from 100 mg of plant material was flash-frozen upon harvesting and ground. The samples were homogenized in 1 mL of TRIzolTM (ThermoFisher, #15596018) using Silamat S6 (Ivoclar Vivadent). RNA was extracted according to manufacturers instructions.Total RNA: For small RNA extraction from total RNA and TraPR input fraction, 10g of total RNA (as described above) or RNA extracted as described above from TraPR inputs was separated on a 15% denaturing polyacrylamide-urea gel. For size reference, 20l of microRNA marker (New England Biolabs, #N2102S) were loaded in the same gel. Following gel staining with peqGreen (Peqlab, #732-3196), and using the marker as reference, small RNAs ranging from 17-to-25nt were excised and recovered using the ZR small-RNA PAGE Recovery Kit (Zymo Research, #R1070).TraPR: Purifucation of Argonaute-loaded small RNAs was performed according to the protocol described in  DOI: 10.1093/nar/gkaa472. Libraries were prepared according to Hayashi et al., 2016 (DOI: 10.1038/nature20162).