GSM7034430: iMEF EV, 4sU, untreated, biol repl 3; Mus musculus; RNA-Seq
Samples were lysed in TRIzol and total RNA was extracted using the DirectZOL Miniprep kit (Zymo Research). For SLAM conversion, 2 µg of total RNA was mixed with 20 mM Iodacetamide (Thermo Fisher Scientific) and incubated at 50°C shaking for 30 min in the dark. The reaction was stopped by mixing with 100 µM dithiothreitol, and converted RNA was purified using the RNeasy Micro Kit (Qiagen). 100 ng of converted RNA was used for constructing the libraries. RNA libraries for SLAM-seq were prepared using the NEBNext® Ultra II Directional RNA Library Prep Kit for Illumina with the NEBNext PolyA mRNA magnetic isolation module (both New Englang Biolabs) and SPRIselect beads (Beckman Coulter) according to the manufacturer's protocols.