|
| Status |
Public on Dec 31, 2014 |
| Title |
EC SA001 day14 batch 2.0 |
| Sample type |
RNA |
| |
|
| Source name |
Human Embryonic Stem Cell line SA001 from Cellartis AB differentiated into Endothelial Cells; batch 2.0
|
| Organism |
Homo sapiens |
| Characteristics |
phenotype: normal-EC_day14
|
| Treatment protocol |
cells were flash frozen and stored at -80°C until RNA extraction
|
| Growth protocol |
Primary endothelial and vascular smooth muscle cells were cultured at 37°C, 5% CO2 and 95% humidity in EGM2 medium (Lonza) until they reached a confluency of maximal 80%.
The Human Embryonic Stem Cell line SA001 (Cellartis AB) was cultured in Priming Medium, consisting N2B27 medium (1:1 mixture of DMEM:F12 (1:1) with Glutamax (Invitrogen) and Neurobasal media supplemented with N2 and B27 (all Invitrogen)) with 1 μM CP21R7 (Roche) and 25 ng/ml BMP4 (R&D Systems)
For differentiation into VSMCs, Human Embryonic Stem Cell line SA001 was cultured in VSMC Induction Medium consisting of N2B27 medium supplemented with 10ng/ml PDGF-BB and 2ng/ml Activin
For differentiation into VSMCs, Human Embryonic Stem Cell line SA001 was cultured in VSMC Expansion Medium consisting of N2B27 with 2ug/ml Heparin and 2ng/ml ActivinA
For differentiation into ECs, Human Embryonic Stem Cell line SA001 was cultured in EC Induction Medium consisting of StemPro-34 SFM medium (Invitrogen) supplemented with 200ng/ml VEGF (PeproTech) and 2μM Forskolin (Sigma-Aldrich)
For differentiation into ECs, Human Embryonic Stem Cell line SA001 was cultured in EC Expansion Medium consisting of StemPro-34 SFM supplemented with 50 ng/ml VEGF
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy Mini kit (Qiagen Inc., Valencia, CA, USA) . DNA was removed by on-column DNase digestion with the RNase-Free DNase set (Qiagen). RNA quality and concentration was determined by analysis with an Agilent 2100 bioanalyzer (Agilent Technologies Inc., Santa Clara, CA, USA).
|
| Label |
Cy3
|
| Label protocol |
Labeling was performed using 5 ug of total RNA input following the standard operating protocol of NimbleGen Systems Inc., Madison, WI USA. See www.nimblegen.com.
|
| |
|
| Hybridization protocol |
Hybridization was performed following the standard operating protocol of NimbleGen Systems Inc., Madison, WI USA. See www.nimblegen.com.
|
| Scan protocol |
Scanning was performed on a NimbleGen MS200 Microarray Scanner at 2 μm pixel resolution following the standard operating protocol of NimbleGen Systems Inc., Madison, WI USA. See www.nimblegen.com.
|
| Description |
ECs in vitro differentiated from HESC SA001 enriched with CD144 Microbeads and an autoMACSpro (Miltenyi Biotec) and cultivated in EC Expansion Medium on human fibronectin (Sigma-Aldrich). Total cultivation time starting from HESC plating is 14 days (day 14). It is the third of three wild-type replicates used in this experiment, each from separate cultures.
|
| Data processing |
The raw data (.pair file) was subjected to background correction, quantile normalization (Bolstad et al. Bioinformatics 19(2):185), and RMA (Robust Multi-Array Analysis; Irizarry et al. Biostatistics 4(2):249) as implemented in the NimbleScan software package, version 2.6 (Roche NimbleGen, Inc.).
|
| |
|
| Submission date |
Jul 11, 2014 |
| Last update date |
Dec 31, 2014 |
| Contact name |
Tobias Heckel |
| E-mail(s) |
tobias-heckel@t-online.de
|
| Organization name |
Roche
|
| Department |
pRED
|
| Lab |
Safety Genetics & Genomics
|
| Street address |
Grenzacherstr. 124
|
| City |
Basel |
| ZIP/Postal code |
CH-4070 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL16025 |
| Series (1) |
| GSE59326 |
Expression profiling in primary and stem-cell derived Human Endothelial and Vascular Smooth Muscle Cells |
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