JP147 3
Homo sapiens
Protocols: Samples were collected by aspirating stimulation medium and washing each well with PBS once. Thereafter cells were lysed using the lysis buffer from  E.Z.N.A. Total RNA Kit I (Omega Bio-tek) according to the manufacturer's instructions. HEK293T cells were seeded at a density of 3 · 106 cells per well in 12-well plates and complete culture medium. After 24 h, medium was changed to DMEM high glucose supplemented with sodium pyruvate (1 mM), penicillin-streptomycin (100 U/mL and 100 µg/mL, respectively) and 0.2% FCS for synchronization over 24 h. Medium was changed again to DMEM high glucose supplemented with sodium pyruvate (1 mM), penicillin-streptomycin (100 U/mL and 100 µg/mL, respectively) and 0.2% FCS, additionally containing either test compounds  (1 µM) in 0.1% DMSO or 0.1% DMSO. After 17 h, cells were harvested the cells.  Total RNA was isolated using E.Z.N.A. Total RNA Kit I (Omega Bio-tek) according to the manufacturer's instructions. RNA concentration and purity were assessed using a NanoDrop One UV–vis spectrophotometer (Thermo Fisher Scientific) at 260/280 nm. Novoegene repeated the isolation using poly-T oligo-attached magnetic beads. After fragmentation, the first strand cDNA was synthesized using random hexamer primers, followed by the second strand cDNA synthesis using either dUTP for directional library or dTTP for non-directional library. For the non-directional library, it was ready after end repair, A-tailing, adapter ligation, size selection, amplification, and purification. For the directional library, it was ready after end repair, A-tailing, adapter ligation, size selection, USER enzyme digestion, amplification, and purification. The library was checked with Qubit and real-time PCR for quantification and bioanalyzer for size distribution detection.
ENA first public
2025-05-01
INSDC center name
Chair for Pharmaceutical and Medicinal Chemistry, Prof. Dr. Daniel Merk, Department Pharmazie, Ludwig-Maximilians-Universität München, Munich, Germany
INSDC status
public
Submitter Id
E-MTAB-15058:JP147 3
broker name
ArrayExpress
cell line
HEK293T
cell type
epithelial
collection date
not collected
common name
human
developmental stage
embryo stage
disease
normal
genotype
wild type genotype
geographic location (country and/or sea)
not collected
isolate
not applicable
organism part
adrenal cortex
sample name
E-MTAB-15058:JP147 3
scientific_name
Homo sapiens