title: GSM4263927: RPL15_overexpressing_BRx142_batch1; Homo sapiens; OTHER
library_construction_protocol: Ribosome profiling was performed as previously described (PUBMED ID 22836135). Briefly, 10 million RPL15- or control-expressing Brx-142 cells were treated with 0.1 mg/ml cyclohexamide for 1 minute, washed with cold PBS containing cycloheximide and lysed. A range of RNase I (Thermofisher) concentrations was tested, and an optimal concentration was chosen that did not lead to degradation of the ribosome protected fragments. RNase I (Thermofisher) treatment was performed and the monosomes were isolated by gel filtration MicroSpin S-400 HR Columns (GE Healthcare). After RNA extraction using RNA Clean & Concentrator-25 kit (Zymo Research), rRNA was depleted (Ribo-Zero Gold rRNA Removal Kit, Illumina), and ribosome-protected fragments were purified by PAGE. The fragments were end- repaired with PNK (NEB). Libraries were prepared using a TruSeq small RNA Library Prep Kit (Illumina) and sequenced on a NextSeq 500 Illumina (76 bp single-end reads).
attributes.tag: GEO Accession
attributes.value: GSM4263927