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| Status |
Public on May 15, 2015 |
| Title |
H1 Day 2 |
| Sample type |
SRA |
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| Source name |
human embryonic stem cells (H1)
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| Organism |
Homo sapiens |
| Characteristics |
cell type: H1
differentiation time: 2 days
differentiation stage: mesoderm committed cells
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| Treatment protocol |
Pluripotent stem cells were differentiated as previously described (Kurian et al, 2012). Briefly, undifferentiated human ES/iPS cells were freshly split on to matrigel-coated plates, making sure the sub-colonies were of small size (~300-500 cells /colony). Cells were differentiated using the chemically defined mesodermal induction media (MIM) (DMEM:F12, 15mg/ml stem cell grade BSA (MP biomedicals), 17.5µg/ml Human Insulin, (SAFC bioscience), 275 µg/ml Human holo transferrin (Sigma Aldrich), 20ng/ml bFGF (Hymanzyme), 50ng/ml Human VEGF-165 aa (Humanzyme), 25ng/ml human BMP4 (Stemgent), 450µM monothioglycerol (Sigma Aldrich), 2.25mM each L-Glutamine and Nonn-essential amino acids (Invitrogen).
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| Growth protocol |
Human ES cells, H1 (WA1, WiCell), (passage 25-40) were cultured in chemically defined hES/hiPS growth media, mTeSR on growth factor reduced matrigel (BD biosciences) coated plates. Briefly, 70-80% confluent hES/hiPS cells were treated with dispase (Invitrogen) for 7 minutes at 37¡C and the colonies were dispersed to small clusters and lifted carefully using a 5ml glass pipette, at a ratio of ~1:6. Human Umbilical Vein Endothelial Cells (HUVEC) were purchased from Promocell and cultivated in EBM medium supplemented with EGM singlequots (Lonza). iPS/ES-derived endothelial cells were cultivated in EGM-2 bullet kit (Lonza). All the endothelial cells were grown on collagen I coated plates (BD biosciences). All cell lines were maintained in an incubator (37¡C, 5% CO2) with media changes every day (hES/hiPS) or every second day (HUVEC).
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA from roughly 10x106 cells was isolated using TRIzol reagent (Invitrogen). Intact total RNA samples were treated with DNase1. All the RNA samples met the following RNA quality- OD260/280 = 2-2.2; OD260/230 ³ 2.0; 28S:18S > 1.0, RIN>7.
TruSeq Stranded Total RNA with Ribo-Zero Human kit (Illumina) was used to remove ribosomal RNA and prepare strand specific paired end RNA seq libraries.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
Total RNA (rRNA depleted with ribozero)
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| Data processing |
Illumina Casava1.7 software used for basecalling.
Reads were aligned to the human genome (hg19) with STAR (v2.2.0c) with default parameters. Only uniquely alignable reads were used for downstream analysis
Reads Per Kilobase of exon per milion mapped reads were calculated using HOMER (http://homer.salk.edu/homer/) across the entire gene (both introns and exons)
Genome_build: hg19
Supplementary_files_format_and_content: Tab delimited text file, Column Def: 1-ensembl ID, 2-chr, 3-gene start, 4-gene end, 5-strand, 6-gene length, 7-copies in genome, 8-gene name, 9-chr band, 10-gene type, 11-status, 12-aliases, 13-18: RPKM values
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| Submission date |
Feb 13, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Christopher Benner |
| E-mail(s) |
cbenner@ucsd.edu
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| Organization name |
University of California, San Diego (UCSD)
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| Department |
Medicine
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| Street address |
9500 Gilman Dr. MC 0640
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| City |
La Jolla |
| State/province |
California |
| ZIP/Postal code |
92093-0640 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (2) |
| GSE54968 |
Transcriptomic analysis reveals novel long non-coding RNAs critical for vertebrate development [RNA-Seq] |
| GSE54969 |
Transcriptomic analysis reveals novel long non-coding RNAs critical for vertebrate development |
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| Relations |
| BioSample |
SAMN02641253 |
| SRA |
SRX470242 |
| Supplementary data files not provided |
SRA Run Selector |
| Processed data are available on Series record |
| Raw data are available in SRA |
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