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Sample GSM3003462 Query DataSets for GSM3003462
Status Public on Apr 30, 2019
Title Day5_CPC_Baf60c_KD_RNA_seq_Rep2
Sample type SRA
 
Source name Cardiac Progenitors
Organism Mus musculus
Characteristics cell line: Baf60c KD E14 Tg(Nkx2-5-EmGFP) ES cells
cell type: Embryonic stem cells (ESC)
sample description: RNA-Seq of Baf60c KD ESC-derived cardiac progenitor cells
Growth protocol Baf60c KD E14 Tg(Nkx2-5–EmGFP) ES cells were maintained on mitomycin treated MEF in Knockout DMEM medium containing 4.5 mg/ml D-glucose, supplemented with 10% serum replacement, 2 mM L-Glutamine, 0.1 mM 2-mercaptoethanol, 1 mM sodium pyruvate, and 1,000 U/ml of leukemia inhibitory factor. For directed cardiomyocyte differentiation, ES cells were grown on gelatin in Neurobasal medium: DMEM/F12 (1:1) medium supplemented with 2,000 U/ml LIF and 10 ng/ml BMP4 for 2 days and differentiated by aggregation in low attachment bacterial dishes at a cell density of 75000 cells/ml in IMDM: F12 medium (3:1). After 48h aggregates were dissociated and re-aggregated in the presence of 5 ng/ml Activin A, 5ng/ml VEGF and 0.2ng/ml BMP4. 40h following the second aggregation, aggregates were dissociated and plated as a monolayer in Stempro-34 medium supplemented with Stempro34 nutrient supplement, L-Ascorbic Acid and 5ng/ml VEGF, 10ng/ml bFGF, and 25ng/ml FGF10 growth factors. Nkx2.5-GFP positive CPC cells were FACS sorted for RNA extraction.
Extracted molecule polyA RNA
Extraction protocol Control and Baf60c KD Nkx2.5-GFP ES cell lines were differentiated into cardiac progenitors. Nkx2.5-GFP positive cells from two different control and Baf60 KD ESCs pools were sorted and RNA was isolated using RNeasy microarray kit (Qiagen #73304). The integrity of RNA was assessed on Bioanalyzer 2100 (Agilent). For RNA-Seq of control and Baf60c KD CPCs, 1 µg RNA was used as input for Truseq Stranded mRNA library preparation (Illumina). Sequencing was performed on NextSeq500 (Illumina) using V2 chemistry.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2500
 
Data processing RNA-Seq reads were mapped to the mm9 reference genome using STAR (−alignIntronMin 20 −alignIntronMax 500000). Samples were quantified using analyzeRepeats.pl (mm9 –count exons –strand both –noadj). Differential expression (fold change >1.5; log2 fold change <−0.58, >0.58; p-value < 0.05) was quantified and normalized using DESEq2. Reads per kilobase per millions mapped (rpkm) was determined using rpkm.default from EdgeR. The KNIME 2.9.1 platform (Konstanz, Germany) was used to filter differentially regulated genes.
Genome_build: mm9
Supplementary_files_format_and_content: Day5_CPC_Baf60c_quanty_transcripts_diff_expr.xls
 
Submission date Feb 13, 2018
Last update date May 03, 2019
Contact name Gergana Dobreva
E-mail(s) Gergana.Dobreva@medma.uni-heidelberg.de
Organization name Medical Faculty Mannheim/University of Heidelberg
Department Cardiovascular Genomics and Epigenomics
Lab AG Dobreva
Street address Ludolf-Krehl-Str. 7-11
City Mannheim
State/province Baden Württemberg
ZIP/Postal code 68167
Country Germany
 
Platform ID GPL17021
Series (2)
GSE80382 Pioneering function of Isl1 in the epigenetic control of cardiomyocyte cell fate [Baf60c_RNA-seq]
GSE80383 Pioneering function of Isl1 in the epigenetic control of cardiomyocyte cell fate
Relations
BioSample SAMN08533378
SRA SRX3697299

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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