biostudies-arrayexpressUnknownTranscriptomicsGenomicsProteomicsAlessia BuratinIllumina NovaSeq 6000RNA-seq of coding RNAHomo sapiensHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-16337We performed RNA sequencing to characterize AGO2-bound and total RNA fractions in four T-cell acute lymphoblastic leukemia (T-ALL) cell lines: JURKAT, ALL-SIL, DND-41 and LOUCY. Libraries were generated from total RNA (TOT) and AGO2-IP RNA samples using the TruSeq Stranded Total RNA kit with Ribo-Zero Human/Mouse/Rat (ribo-depletion protocol; fragmentation at 90 °C for 2 minutes [doi:10.1016/j.ygeno.2021.10.018]). Sequencing was carried out on an Illumina NovaSeq 6000 platform (2×150 bp paired-end, ~150 million read pairs per sample).biostudies-arrayexpressSample Treatment - AGO2 immunoprecipitation (AGO2-IP) was performed as described previously with some minor adaptations [doi: 10.1093/nar/gkp715]. Briefly, cell lysates were incubated with EZview Red Protein G Affinity Gel (Sigma-Aldrich, St. Louis, MO, USA) coated with anti-AGO2 antibody (Clone 2E12-1C9, Abnova, Taipei, Taiwan) at 4°C overnight. Anti-IgG antibody was used as a negative control (Merck, Darmstadt, Germany). 10% of input, AGO2-IP and flow-through fractions were collected to test the effectiveness of the AGO2 RNA IP procedure via AGO2 immunoblotting and RT-qPCR for selected miRNAs. The remaining AGO2-IP fraction was further used for RNA isolation and sequencing.Nucleic Acid Extraction - The miRNeasy micro RNA isolation kit (Qiagen, Hilden, Germany) was used for the extraction of total RNA (TOT). RNA isolates were DNase-treated and purified with the use of RNA Clean and Concentrator Kit (Zymo Research, Irvine, CA, USA). AGO2-IP RNA was isolated directly using the RNA Clean and Concentrator Kit. RNA concentration was measured with Quantus Fluorometer (Promega, Madison, WI, USA) using Qubit HS RNA Assay Kit (Thermo Fisher Scientific). RNA integrity was determined with 4200 Tapestation using High Sensitivity RNA ScreenTape (Agilent Technologies, Santa Clara, CA, USA).Library Construction - Libraries were constructed from the total (TOT) and AGO2-IP RNA fraction samples in the 4 T-ALL cell lines, using TruSeq Stranded Total RNA with Ribo-Zero Human/Mouse/Rat protocol, based on ribo-depletion. Fragmentation was performed at 90′C for 2 minutes, as described previously [https://doi.org/10.1016/j.ygeno.2021.10.018].Growth Protocol - Cells were cultured under standard conditions in RPMI-1640 medium (Thermo Fisher Scientific, Waltham, MA, USA) with 10% (JURKAT, LOUCY, DND-41) or 20% (ALL-SIL) of fetal bovine serum (Thermo Fisher Scientific).Sequencing - The libraries were sequenced on an Illumina NovaSeq6000 platform, using the following settings: 2x150 PE (paired end sequencing with 150 nt reads), read depth of coverage: 150 million read pairs/sample.Sample Collection - For each immunoprecipitation reaction, 3 × 10^7 cells were used. 10% of input (3 x 10^6) was collected as total RNA (TOT) fraction.OrganizationMINSEQE ScoreAssays and DataMAGE-TAB FilesAlessia BuratinfalseAGO2 immunoprecipitation RNA-seq in T-ALL cell linesWe performed RNA sequencing to characterize AGO2-bound and total RNA fractions in four T-cell acute lymphoblastic leukemia (T-ALL) cell lines: JURKAT, ALL-SIL, DND-41 and LOUCY. Libraries were generated from total RNA (TOT) and AGO2-IP RNA samples using the TruSeq Stranded Total RNA kit with Ribo-Zero Human/Mouse/Rat (ribo-depletion protocol; fragmentation at 90 °C for 2 minutes [doi:10.1016/j.ygeno.2021.10.018]). Sequencing was carried out on an Illumina NovaSeq 6000 platform (2×150 bp paired-end, ~150 million read pairs per sample).2025-12-29T00:00:00Z2025-12-29T02:02:03.98Z2025-12-02T15:13:44.108ZE-MTAB-16337ERP185968EFO_0002944EFO_0004170EFO_0003789EFO_0005518EFO_0003738EFO_0004184EFO_0003969