title: Sample 14
organism: Homo sapiens
description: Protocols: hiPSC-derived endothelial cells were generated using a previously published protocol. hiPSCs from three different lines r-iPSC1J, ChiPS22 (TakaraBio) and C9 were seeded at 15,500 17,000 and 17,000 cells/cm2, respectively, in DEF-CS 24 hours before the start of differentiation. Differentiation was initiated by changing the medium to Essential 6 (E6) media (Gibco). Cultures were given fresh E6 daily, for four days. On day four, E6 was changed to endothelial cell media [EC-media, EC-SFM (Life Technologies), 1 % platelet-poor serum (Alfa Aesar), 10 µM retinoic acid (Sigma Aldrich) and 20 ng/ml bFGF (Peprotech)]. On day six cells were passaged at 1,000,000 cells/cm2 onto collagen (400 µg/ml, Sigma Aldrich)/fibronectin (100µg/ml Sigma Aldrich)-coated 24-well 0.4 µm pore polyester membrane Transwell inserts (Corning) or 6-well plates (Corning). After 24 hours, on day seven, the media was changed to EC-media without bFGF and retinoic acid. The next day, on day 8, cells were dissociated and cryopreserved in 90% fetal bovine serum (Gibco), 10 % DMSO (Sigma Aldrich) or seeded in OrganoPlates at 20,000 cells/µl. Immortalized human BEC CMEC/D3 (Merck) were maintained according to the vendor's instructions on collagen I (Sigma) coated plates in EndoGro (Millipore) media. Human umbilical vein endothelial cells (HUVECs) (Lonza) were maintained according to the vendors instructions in EGM2 growth media (Lonza). Samples for RNA sequencing were collected after 48h after confluence was reached. Fifteen samples from Transwell (2D) and fifteen from OrganoPlate (3D) were taken, each including three replicates from C9, r-iPSC1J, ChiPS22, HUVECs and CMEC, respectively. Approximately 1µg RNA per sample was collected and RNA was purified using the RNeasy Mini Kit (Qiagen) with DNase treatment according to the manufacturer's instructions. Sequencing library construction was performed using Illumina TruSeq stranded mRNA with poly-A selection.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2021-02-01T04:05:23Z
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2020-02-11T17:18:21Z
attributes.tag: External Id
attributes.value: SAMEA6535965
attributes.tag: INSDC center name
attributes.value: School of Bioscience, University of Skovde, Sweden
attributes.tag: INSDC first public
attributes.value: 2021-02-01T04:05:23Z
attributes.tag: INSDC last update
attributes.value: 2020-02-11T17:18:21Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-8791:Sample 14
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell line
attributes.value: r-iPSC1J
attributes.tag: cell type
attributes.value: endothelial cell
attributes.tag: common name
attributes.value: human
attributes.tag: developmental stage
attributes.value: embryo
attributes.tag: disease
attributes.value: normal
attributes.tag: genotype
attributes.value: wild type genotype
attributes.tag: individual
attributes.value: P13674_114
attributes.tag: organism part
attributes.value: skin
attributes.tag: progenitor cell type
attributes.value: induced pluripotent stem cell
attributes.tag: sample name
attributes.value: E-MTAB-8791:Sample 14
attributes.tag: scientific_name
attributes.value: Homo sapiens