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Sample GSM7720585 Query DataSets for GSM7720585
Status Public on Apr 23, 2024
Title HEK293T, WT, groseq, Rep1
Sample type SRA
Source name kidney
Organism Homo sapiens
Characteristics tissue: kidney
cell line: HEK293T
cell type: human embryonic kidney
genotype: wild type
fraction: nascent RNA
Treatment protocol LN-229 cells were transfected in triplicate with 100nM of siRNA oligonucleotides targeting human ZFX (Horizon #L006572000005) or control oligonucleotides (Horizon #D0018101005) using Lipofectamine RNAiMAX (Thermo Fisher #13778150) for 72 hours.
Growth protocol LN-229 and HEK293T were grown using DMEM supplemented with 10% FBS and pen/strep.
Extracted molecule total RNA
Extraction protocol 5x106 cells per nuclear run-on reaction were harvested and incubated in swelling buffer (10 µM Tris-HCl, 2 mM MgCl2, 3 mM CaCl2). Nuclei were isolated with lysis buffer (10 µM Tris-HCl, 2 mM MgCl2, 3 mM CaCl2, 10% glycerol, 1% NP-40). Nuclear run-on was performed at 30°C for 5 min in 10 mM Tris-HCl pH 8, 5 mM MgCl2, 300 mM KCl, 1 mM DTT, 500 µM ATP, 500 µM GTP, 500 µM Br-UTP, 2 µM CTP, 200 U/mL Superase In RNase Inhibitor (Invitrogen #AM2696), and 1% sarkosyl. Nuclear RNA was isolated with Trizol (Invitrogen #15596026). RNA was purified with Micro Bio-Spin P-30 Gel Columns (Bio-Rad # 7326250), fragmented with RNA Fragmentation Kit (Invitrogen # AM8740), and treated with 15 units mRNA decapping enzyme (NEB # M0608S) and 30 units T4 PNK (NEB # M0201S). RNA immunoprecipitation was performed 3 times with anti-BrdU-conjugated agarose beads (Santa Cruz Biotechnologies # sc-32323 AC).
Library preparation was performed with TruSeq Small RNA Library Preparation Kit (Illumina # RS-200-0012 following the manufacturer’s protocol.
Library strategy OTHER
Library source transcriptomic
Library selection other
Instrument model Illumina NovaSeq 6000
Data processing Reads with Illumina small RNA adapter sequences were trimmed using Trimgalore
Reads were aligned to hg38 using Tophat
Normalization was done across samples using an equal number of uniquely mapped reads
Assembly: hg38
Supplementary files format and content: bigwig file with total mapped reads
Library strategy: GRO-seq
Submission date Aug 21, 2023
Last update date Apr 23, 2024
Contact name Peggy J Farnham
Organization name University of Southern California
Department Biochemistry and Molecular Medicine
Street address 1450 Biggy St, NRT6514
City Los Angeles
State/province CA
ZIP/Postal code 90033
Country USA
Platform ID GPL24676
Series (2)
GSE241281 Reduction of ZFX levels decreases histone H4 acetylation and increases Pol2 pausing at target promoters [GRO-seq]
GSE241284 Reduction of ZFX levels decreases histone H4 acetylation and increases Pol2 pausing at target promoters
BioSample SAMN37106835
SRA SRX21447431

Supplementary file Size Download File type/resource
GSM7720585_USC_GRO001_293t_rep1_groseq_minus.bigwig 86.0 Mb (ftp)(http) BIGWIG
GSM7720585_USC_GRO001_293t_rep1_groseq_plus.bigwig 89.7 Mb (ftp)(http) BIGWIG
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Raw data are available in SRA

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