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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Aug 01, 2018 |
| Title |
3367-HJ-0004 |
| Sample type |
SRA |
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| Source name |
Spleen
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| Organism |
Mus musculus |
| Characteristics |
cell types: Purified B220+ cells
genotype: WT
treatment: none
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| Growth protocol |
P493-6 cells were cultured in RPMI 1640 medium with 10% fetal bovine serum (FBS).
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNAs were isolated using RNeasy Plus Mini Kit (Qiagen). ERCC RNA Spike-In Mix (Ambion, 4456740) was used following instructions of the product. The amount of spike-in added was calibrated to the RNA yield to ensure the spike-in signal was in the appropriate dynamic range. Specifically, 2μl of a 1:200 diluted ERCC Spike-In Mix 1 was added to one of the samples that had a total RNA of 465ng. Between 1.8μl to 5.4μl of 1:200 diluted ERCC Spike-In Mix 1 were added to other samples (between 240 to 800ng total RNAs) at the same ratio of Spike-in/cell number.
Library preparation and sequencing for RNA and ChIP (and their corresponding input) DNA samples were performed at the Genomic Services Lab at HudsonAlpha Institute for Biotechnology (Huntsville, AL). Briefly, the quality of the total RNA and DNA was assessed using the Agilent 2100 Bioanalyzer. Two rounds of polyA+ selection was performed for RNA samples, followed by conversion to cDNAs. The mRNA library generation kits (Agilent, Santa Clara, CA) and TruSeq ChIP Sample Prep Kit (Illumina) were used generate sequencing libraries per manufacturer’s instructions. The indexed DNA libraries were quantitated using qPCR in a Roche LightCycler 480 with the Kapa Biosystems kit for library quantitation (Kapa Biosystems, Woburn, MA) prior to cluster generation. Clusters were generated to yield approximately 725 K to 825 K clusters/mm2. Cluster density and quality were determined during the run after the first base addition parameters were assessed.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Basecalls performed using CASAVA version 1.8.4
For RNA-seq, all the reads were mapped to the mouse reference genome (GRCm38/mm10) using TopHat (v2.0.13). The alignment was guided using a Gene Transfer File (GTF version GRCm38.85). Low quality mapped reads (MQ<30) were removed from the analysis. Read count tables were generated using HTSeq (v.0.6.0) based on the Ensembl gene annotation file (Ensembl GTF version GRCm38.85) (--mode=union --stranded=no --minaqual=30 --idattr=gene_name –type=exon). All the read count tables were then normalized based on their ERCC RNA Spike-In Mix size factors calculated using the DESeq R package (v.3.0), and Deferential Expression (DE) analyses were performed using DESeq (v3.0). All of the downstream statistical analyses and generating plots were performed in R (v3.1.1). To generate the heat map, upregulated and downregulated genes were clustered separately using R dist function in Euclidian mode and then merged to maintain the clustered order of the genes. The values in the matrix were scaled based on columns and not rows using heatmap.2 R function to make the two different data comparable in a single heat map.
Genome_build: hg19, mm10
Supplementary_files_format_and_content: BigWig and TDF (coverage for mapped reads), count tables (read counts for genomic features)
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| Submission date |
Jul 25, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Hao Jiang |
| E-mail(s) |
haojiang@uab.edu
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| Organization name |
University of Alabama at Birmingham
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| Department |
Biochemistry and Molecular Genetics
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| Street address |
1825 University Blvd
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| City |
Birmingham |
| State/province |
AL |
| ZIP/Postal code |
35294 |
| Country |
USA |
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| Platform ID |
GPL17021 |
| Series (2) |
| GSE101853 |
Dpy30 regulates Myc binding to targets and Myc-driven tumorigenesis (ChIP-seq, RNA-seq) |
| GSE101854 |
Dpy30 regulates Myc binding to targets and Myc-driven tumorigenesis |
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| Relations |
| BioSample |
SAMN07413937 |
| SRA |
SRX3033542 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2717257_genes_HJ0004.txt.gz |
147.0 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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