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Sample GSM4487557 Query DataSets for GSM4487557
Status Public on Apr 19, 2022
Title Sample 8: P1.T.M.2
Sample type SRA
 
Source name Primary CLL cells
Organism Homo sapiens
Characteristics patient: P1
disease state: Chronic Lymphocytic Leukemia
treatment: siZAP70
replicate: Rep 2
Treatment protocol For samples with anti-IgM treatment, primary CLL cells were co-cultured with EL08 stromal cells and treat with anti-IgM 4ug/1x10^6 CLL cells.
Growth protocol Primary CLL cells were transfected with NSC or ZAP-70 siRNA and co-cultured with EL08 stromal cells in RPMI 1640 medium for totally 7 days.
Extracted molecule total RNA
Extraction protocol Primary CLL cells were harvested after positivley selected by anti-CD19 beads and total RNA was extracted using the standard protocol from GenElute Mammalian Total RNA Miniprep Kit (Sigma/RTN70).
NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina (NEB #7420) was used with 100ng of total RNA for the construction of sequencing libraries. mRNA was isolated and purified by using NEBNext Poly (A) mRNA Magnetic Isolation Module (NEB #7490) before cDNA synthesis.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 4000
 
Data processing The quality of the data was assessed using FastQC, MultiQC, PCAs, Jaccard Similarity Indices and MA plots
the reads were aligned using STAR v2.7.3a (parameters --outSAMtype BAM SortedByCoordinate --outReadsUnmapped Fastx --readFilesCommand gunzip -c)
Protein-coding gene quantification was performed using Subread featureCounts v2.0.0 (default parameters)
The raw expression levels were normalized with edgeR v3.26.8. After normalization, a noise filtering step was applied i.e. for each gene, if the maximum normalised expression level was <10 then the gene was excluded from the downstream analysis.
Statistical testing for differential expression of genes was performed using edgeR, with multiple-testing correction based on the Benjamini-Hochberg method. DE Genes were called on adj p-value < 0.05, and |log2(FC)| > 0.5.
The enrichment analysis was performed using g:profiler (Reimand et al., Nucleic Acids Research, 2016)
Genome_build: GRCh38.p13
Supplementary_files_format_and_content: raw.csv contains raw gene counts; normalized.csv contains edgeR-normalized expression levels after the application of the noise filter
 
Submission date Apr 21, 2020
Last update date Apr 19, 2022
Contact name Irina Mohorianu
E-mail(s) data-submissions@stemcells.cam.ac.uk
Organization name University of Cambridge
Department Wellcome-MRC Cambridge Stem Cell Institute
Street address Puddicombe Way
City Cambridge
ZIP/Postal code CB2 0AW
Country United Kingdom
 
Platform ID GPL20301
Series (2)
GSE149020 Next generation sequencing quantitatively analyzes transcriptomes of primary CLL cells tranfected with non-specific control (NSC) or ZAP-70 siRNA (-anti-IgM dataset)
GSE149022 Next generation sequencing quantitatively analyzes transcriptomes of primary CLL cells tranfected with non-specific control (NSC) or ZAP-70 siRNA
Relations
BioSample SAMN14652399
SRA SRX8148311

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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