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Status |
Public on Dec 01, 2016 |
Title |
RNA shKdm5ABC 4 hour exp 1 |
Sample type |
SRA |
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Source name |
3T3-L1_RNA shKdm5ABC 4 hour
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Organism |
Mus musculus |
Characteristics |
cell line: 3T3-L1 cell type: Preadipocyte cell line transduced with: shKdm5ABC time point: 4 hours post induction molecule subtype: polyA-enriched mRNA
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Treatment protocol |
Two days post confluency (day 0), 3T3-L1 cells were induced to differentiate using a standard cocktail of adipogenic inducers (dexamethasone, insulin, 3-isobutyl-1-methylxanthine, and fetal bovine serum). Two days post induction (day 2), medium was changed to DMEM supplemented with fetal bovine serum and insulin. Cells were maintained in DMEM supplemented with fetal bovine serum for the remainder of the differentiation. For knock down experiments, pre-confluent 3T3-L1 cells were transduced 4 days before induction of differentiation by incubation for 24 hours at 37°C with medium containing lentiviral constructs (shKDM5A/B/C or control shLuc) and 6 μg/mL polybrene (Sigma). After 24 hours the medium was changed to normal growth medium.
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Extracted molecule |
polyA RNA |
Extraction protocol |
Total RNA from 3T3-L1 cells was purified using TRIzol (Invitrogen) according to the manufacturer’s instructions. RNA was treated and the integrity of the RNA was confirmed using a Bioanalyzer 2100 (Agilent Technologies). 4 ug of total RNA was polyA-selected, and subsequently subjected to fragmentation and cDNA-synthesis using the TruSeq RNA Sample Prep kit v2 (Illumina). cDNA-seq libraries were constructed according to the manufacturer's instructions (Illumina) as described in (Nielsen R, Mandrup S, Methods in Enzymology 2014, Vol. 537, pp. 261-279).
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Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 1500 |
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Description |
processed data file: RNA_all_genes_log2_tags_kb.txt
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Data processing |
RNA reads were aligned to the mouse reference genome (version mm9) using Bowtie2 (version 0.12.8) (Langmead and Salzberg 2012, Nat Met 9: 357-359) with standard parameters. Splice-junction reads were handled by creation of a pseudo-splice genome, similar to the strategy utilized in RSEQTools (Habegger et al. 2011, Bioinformatics 27: 281-283). Reads were filtered post-alignment for a MAPQ score greater than or equal to 30. The number of exon reads for all RefSeq genes were counted using Subread (Liao et al. 2013, Nucleic Acids Res 41: e108) and differential expression analyses were performed using DESeq2 (FDR <0.05, paired analysis) (Love et al., 2014, Genome Bio. 15: 550). Genome_build: mm9 Supplementary_files_format_and_content: H3K4me3_promoter_peaks_tags.txt: Table containing promoter H3K4me3 binding sites, including normalized tag counts for H3K4me3 libraries KDM5A_all_peaks_tags.txt: Table containing all identified KDM5A binding sites, including normalized tag counts for KDM5A libraries KDM5C_all_peaks_tags.txt: Table containing all identified KDM5C binding sites, including normalized tag counts for KDM5C libraries RNA_all_genes_log2_tags_kb.txt: Table containing gene expression data for all RNA-seq libraries (reported as log2(tags/kb))
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Submission date |
Jul 14, 2016 |
Last update date |
May 15, 2019 |
Contact name |
Ann-Sofie Bøgh Brier |
E-mail(s) |
a.boegh@me.com
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Phone |
+4522969284
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Organization name |
University of Southern Denmark, Odense
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Department |
Department of Biochemistry and Molecular Biology
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Lab |
Mandrup lab
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Street address |
Campusvej 55
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City |
Odense M |
ZIP/Postal code |
5230 |
Country |
Denmark |
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Platform ID |
GPL18480 |
Series (1) |
GSE84410 |
The KDM5 family is required for activation of pro-proliferative cell cycle genes during adipocyte differentiation |
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Relations |
BioSample |
SAMN05391988 |
SRA |
SRX1949124 |
Supplementary data files not provided |
SRA Run Selector |
Raw data are available in SRA |
Processed data are available on Series record |
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