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Sample GSM7329827 Query DataSets for GSM7329827
Status Public on Sep 11, 2023
Title RNA-Seq K562 RED rep2
Sample type SRA
 
Source name blood
Organism Homo sapiens
Characteristics tissue: blood
cell line: K562
disease state: erythroleukemia
treatment: erythroid differentiation
agent: hemin
Treatment protocol K562 cells incubated in the presence of 50 μM hemin (neoFroxx, Germany) for 108 h. At this concentration, the drug did not affect cell proliferation.
Growth protocol The cells were drown in RPMI 1640 media (PanEco, Russia) supplemented with heat-inactivated fetal calf serum (HyClone, USA), 2 mM glutamine, 250 u/ml penicillin and 250 μg/ml streptomycin (PanEco, Russia) at 37 °C in a humidified atmosphere containing 5% CO2. For a treated samples, hemin (50 μM, neoFroxx, Germany) was added to induce the erythroid differentiation and cells were incubated further for 108 h. At this concentration, the drug did not affect cell proliferation.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from cell lysed with Trisol reagent and PureLink RNA Micro Kit (Invitrogen) according to manufacture instruction. RNA Quality was checked with BioAnalyser and RNA 6000 Nano Kit(Agilent). Poly(A)+ RNA was isolated with Dynabeads® mRNA Purification Kit (Ambion).
Poly(A)+ RNA was isolated with Dynabeads® mRNA Purification Kit (Ambion). Illumina library was prepared from poly(A)+RNA with NEBNext® Ultra™II RNA Library Prep Kit for Illumina® (NEB) according to manual. Sequencing was performed on Illumina HiSeq1500 with 50 bp read length.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 1500
 
Description Replicate 2 hemin
RNAseq.RED.forward.bw
RNAseq.RED.reverse.bw
K562.RED.TPM.txt
K562.RED.FPKM.txt
Data processing Base calling and quality control were performed in real time with standard Illumina analysis pipeline using a phiX control.
Sequenced reads were trimmed from low quality and/or short sequences and/or Illumina 3' adapters by trimmomatic v. 0.39 using the following options: LEADING:18 TRAILING:18 SLIDINGWINDOW:4:22 MINLEN:20 ILLUMINACLIP:TruSeq3-SE:2:30:10
Trimmed reads were quantified to H. sapiens hg38 GRCh38 Ensembl v.106 genome with Ensembl genome map v.106 by RSEM 1.3.1 using the following options: --fragment-length-mean 255 --star --calc-ci --ci-memory 30720. This operation was performed for both replicates separately. The results were averaged by an in-house R script.
Trimmed reads were mapped to H.sapiens genome (hg38/GRCh38 Ensembl v.106) by STAR aligner 2.7.5c with hg38 Ensembl genome map v.106 http://ftp.ensembl.org/pub/release-106/gtf/homo_sapiens/Homo_sapiens.GRCh38.106.gtf.gz. Alignment options were the following: --outSAMunmapped Within --outFilterType BySJout --outSAMattributes NH HI AS NM MD --outFilterMultimapNmax 20 --outFilterMismatchNmax 999 --outFilterMismatchNoverLmax 0.04 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --sjdbScore 1 --quantMode TranscriptomeSAM
Alignment files were separated by samtools 1.15 view -F 0x10 to get forward strand alignments and by samtools 1.15 view -f 0x10 to get reverse strand alignments. Then bamCoverage (deepTools 3.5.1) tool was applied to obtain normalized expression profiles: --binSize 10 --effectiveGenomeSize 2913022398 --skipNAs --normalizeUsing RPKM --exactScaling. The obtained profiles were averaged between replicates by WiggleTools v1.2.11 and converted to bigWig format by bedGraphToBigWig tool from UCSC genome tools.
Assembly: Homo sapiens hg38/GRCh38 Ensembl release 106 genome build. Homo sapiens hg38/GRCh38 Ensembl annotation v106 in GTF format.
Supplementary files format and content: bigWig files contain normalized (RPKM, reads per kilobase million) expression signal genomewide for each strand.
Supplementary files format and content: Results files contain quantified expression values in TPM (Transcripts Per Million) and FPKM (Fragments Per Kilobase Million) for each gene per replicate and averaged between replicates.
 
Submission date May 12, 2023
Last update date Sep 11, 2023
Contact name Nickolai Tchurikov
E-mail(s) tchurikov@eimb.ru
Organization name Engelhardt Institute of Molecular Biology
Department Department of Epigenetic Mechanisms of Gene Expression Regulation
Street address 32, Vavilova str.
City Moscow
ZIP/Postal code 119991
Country Russia
 
Platform ID GPL18460
Series (2)
GSE232390 Analysis of gene expression changes in K562 erythroleukemia cells after erythroid differentiation caused by incubation in the presence hemin [RNA-Seq]
GSE232393 K562 erythroleukemia cells after erythroid differentiation caused by incubation in the presence of hemin
Relations
BioSample SAMN35057777
SRA SRX20307690

Supplementary file Size Download File type/resource
GSM7329827_K562.RED.rep2.genes.results.gz 3.4 Mb (ftp)(http) RESULTS
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record
Processed data provided as supplementary file

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