title: GSM8422083: SCAF1AID_4sUSeq_Auxin_Rep2; Homo sapiens; OTHER
library_construction_protocol: Cells were lysed using QIAzol and lysates were spiked with 4sU-treated mouse T-cell lysates. Total RNA was purified with the miRNeasy mini kit, precipitated to reduce volume, and then subject to fragmentation. Fragmented RNA was biotinylated. This was followed by 4sU-RNA pulldown using MyOne streptavidin beads, followed by purification with the RNeasy MinElute cleanup kit. The 4sU-RNA was subject to alkylation using iodoacetamide, and then alkylated-4sU-RNA was sequenced. 4sU-RNA was used for library preparation with NEBNext rRNA Depletion Kit (Human/Mouse/Rat) and NEBNext Ultra II Directional RNA Library Prep kit. The concentration and size distribution of the libraries were determined on a Fragment Analyzer using the NGS Fragment High Sensitivity Analysis Kit (1-6,000 bp; Agilent Technologies). The libraries were sequenced on the NextSeq2000 Illumina platform. Base calling was performed using Illumina's BaseSpace platform.