Homo sapiens
Protocols: Feeder- and serum-free hESC (H9 line; WiCell) culture was previously described (Vallier, 2011). Cells were plated on gelatin- and MEF medium-coated plates, and cultured in chemically defined medium (CDM) containing bovine serum albumin (BSA). CDM was supplemented with 10ng/ml Activin-A and 12ng/ml FGF2 (both from Dr Marko Hyvonen, Dept. of Biochemistry, University of Cambridge). Cells were treated for 2h with fresh culture media containing 10ng/ml Activin-A, or cultured for 2h, 8h, or 32h of with media containing 10uM SB-431542 (SB) instead of Activin-A. Three biological replicates per condition were examined Cellular RNA was extracted using the GenElute Mammalian Total RNA Miniprep Kit and the On-Column DNase I Digestion Set (both from Sigma-Aldrich) following manufacturer's instructions. Polyadenylated (poly-A) purified opposing strand-specific mRNA library libraries were prepared from 200ng of total RNA using the TruSeq Stranded mRNA HT sample preparation kit (Illumina). Samples were individually indexed for pooling using a dual-index strategy. Libraries were quantified both with a Qubit (ThermoFisher Scientific) and by qPCR using the NGS Library. Quantification Kit (KAPA Biosystems). Libraries were then normalized and pooled.
ENA first public
2018-01-04
ENA last update
2016-11-10
External Id
SAMEA4537389
INSDC center alias
Wellcome Trust - Medical Research Council Cambridge Stem Cell Institute, Anne McLaren Laboratory for Regenerative Medicine, Department of Surgery, University of Cambridge
INSDC center name
Wellcome Trust - Medical Research Council Cambridge Stem Cell Institute, Anne McLaren Laboratory for Regenerative Medicine, Department of Surgery, University of Cambridge
INSDC first public
2018-01-04T17:02:34Z
INSDC last update
2016-11-10T08:58:11Z
INSDC status
public
Submitter Id
E-MTAB-5230:Sample 13
broker name
ArrayExpress
cell line
H9
cell type
embryonic stem cell
common name
human
sample name
E-MTAB-5230:Sample 13
sex
female