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Sample GSM4795825 Query DataSets for GSM4795825
Status Public on Oct 30, 2020
Sample type SRA
Source name REH pre-B-ALL cell line
Organism Homo sapiens
Characteristics time: 72 h
construct: Vincristine
treatment: Vincristine
Treatment protocol REH cells were seeded into 6-well plates (0.6 milion/ml concentration) and treated with XRP44X (Sigma-Aldrich) (1 µM), TK216 (MedChemExpress, NJ USA) (800 nM) or DMSO for 72 h
Growth protocol The E/R+ REH cell line (ACC-22, DSMZ, Germany) was maintained in RPMI 1640 (Gibco, Thermo Fisher) supplemented with 10% FBS (Gibco, Thermo Fisher), 2 mM L-glutamine (Gibco, Thermo Fisher), penicillin (100 U/ml), and streptomycin (100 mg/ml) (Sigma-Aldrich).
Extracted molecule polyA RNA
Extraction protocol Viability was checked using Trypan blue with Cellometer Mini Automated Cell Counter (Nexcelom Bioscience) and Dead Cell Removal Kit (#130-090-101, MACS miltenyi Biotech) was used. Viable cells were eluted by rinsing twice with 1 ml binding buffer. Subsequently 0.42-0.5 million cells were methanol fixated according to 10X Genomics Methanol Fixation of Cells for Single Cell RNA Sequencing protocol User guide CG000136 Rev E, using a mix of two RNAse inhibitors (RNase Inhibitor, Thermo Fisher, Carlsbad, CA, USA and RNasin® Plus RNase Inhibitor, Promega, Madison, WI, USA). Samples were stored < 6 weeks before proceeding with Chromium run.
Cells were thawn following the MetOH fixation protocol and processed for loading following the Chromium Single Cell 3´Reagent Kits v3 User guide CG000183 Rev C. REH cells were prepared into pools together with mouse cells (not this study) and human cell singlets separated in data analysis step. Loading concentration of 2100-2200 cells/uL was used. Using 10X GemCode Technology, the poly-A transcripts are barcoded with an Illumina R1 sequence, a 16 bp 10X barcode and a 10 bp Unique Molecular Identifier (UMI). 13 cycles of PCR were used to amplify the cDNA.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina NovaSeq 6000
Description Cell line experiment in REH comparing two drugs targeting ETS-factors at 72 h.Vincristine (1nM) sample.
Data processing Data was aligned with Cell Ranger 3.1.0 to combined human (hg19) and mouse (mm10) genome as reference from 10x Genomics with default parameters.
Cells were filtered with Cell Ranger 3.1.0 following alignment.
The mouse cells were left out by filtering cells which cellranger characterized as mouse or doublet cells or had more than 500 counts assigned to mouse genes.
The mouse genes were filtered out from the count matrix.
Genome_build: hg19
Supplementary_files_format_and_content: barcode annotation: 10x Cell Ranger filtered barcode list; feature annotation: 10x Cell Ranger feature list; count matrix: 10x Cell Ranger filtered UMI count matrix
Submission date Sep 19, 2020
Last update date Nov 17, 2023
Contact name Merja Heinäniemi
Organization name University of Eastern Finland
Department School of Medicine
Street address Yliopistonranta 1 E
City Kuopio
ZIP/Postal code 70211
Country Finland
Platform ID GPL24676
Series (1)
GSE148218 Single cell characterization of arrested B-lymphoid differentiation and leukemic cell states in ETV6-RUNX1-positive pediatric leukemia
BioSample SAMN16221311
SRA SRX9156856

Supplementary file Size Download File type/resource
GSM4795825_REH_Vincristine_barcode_metadata.tsv.gz 21.5 Kb (ftp)(http) TSV
GSM4795825_REH_Vincristine_features.tsv.gz 507.7 Kb (ftp)(http) TSV
GSM4795825_REH_Vincristine_matrix.mtx.gz 26.5 Mb (ftp)(http) MTX
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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