title: Sample 3
organism: Homo sapiens
description: Protocols: At 5 and 7 hpi, LTM-treated and non-treated cells were flash frozen in an ethanol/dry ice bath and lyzed in the presence of 0.36 mM cycloheximide (CHX). HeLa Ohio cells were grown on 150-mm dishes to a confluency of 80% and infected with CVA-13 at an MOI of 10 to ensure synchronous infection. For lactimidomycin (LTM) treated cells, 30 min before the specified time point, cells were treated with 50 mM LTM for 30 min.  Cells were rinsed with 5 ml of ice-cold PBS, the dishes submerged in a surface of dry ice-ethanol bath for 10 s, transferred to dry ice and 400 µl of lysis buffer [20 mM Tris-HCl pH 7.5, 150 mM NaCl, 5 mM MgCl2, 1 mM DTT, 1% Triton X-100, 100 µg/ml cycloheximide and 25 U/ml TURBO DNase (Life Technologies)] dripped on. Cells were scraped extensively to ensure lysis, collected and triturated with a 26-G needle ten times. Lysates were clarified by centrifugation for 20 min at 13,000 g at 4 ºC, the supernatants recovered and stored in liquid nitrogen. The purification of RPFs was performed using Direct-zol RNA MiniPrep kit (Zymo research). The further methodologies employed were based on the original protocols (Ingolia et al., 2012; Irigoyen et al., 2016), except ribosomal RNA contamination was removed using RiboZero Gold rRNA removal kit (Illumina) and library amplicons were constructed using a small RNA cloning strategy (Guo et al., 2010) adapted to Illumina smallRNA v2 to allow multiplexing. Amplicon libraries were deep sequenced using an Illumina NextSeq platform.
attributes.tag: ENA first public
attributes.value: 2025-11-27
attributes.tag: INSDC center name
attributes.value: University of Cambridge
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-14994:Sample 3
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell line
attributes.value: HeLa
attributes.tag: cell type
attributes.value: epithelial cell
attributes.tag: collection date
attributes.value: not collected
attributes.tag: common name
attributes.value: human
attributes.tag: developmental stage
attributes.value: adult
attributes.tag: geographic location (country and/or sea)
attributes.value: not collected
attributes.tag: isolate
attributes.value: not applicable
attributes.tag: organism part
attributes.value: uterine cervix
attributes.tag: replicate
attributes.value: 1
attributes.tag: sample name
attributes.value: E-MTAB-14994:Sample 3
attributes.tag: scientific_name
attributes.value: Homo sapiens
attributes.tag: virus
attributes.value: CVA13