title: RNA sequencing analyses of wild-type and variant 1q human pluripotent stem cells
design: RNA sequencing analyses of wild-type and variant 1q human pluripotent stem cells
library_construction_protocol: Three biological replicates of wild-type and v1q sublines of the H7 and H9hPSC lines from E8/VTN cultures were used for RNA extraction and RNAseq analysis. Prior to RNA isolation, cells were lysed using Buffer RLT (Cat. # 79216; Qiagen) and stored at -80C. Human pluripotent stem cell culture For E8/VTN culture conditions, hPSC were grown on vitronectin (VTN-N) (Cat. # A14700, Life Technologies)-coated flasks (5 µg/ml in Dulbecco's phosphate buffered saline (PBS) without Calcium and Magnesium) in modified E8 medium11 prepared in house, consisting of DMEM/F12 (Cat. # D6421; Sigma-Aldrich) supplemented with 14 µg/l sodium selenium (Cat. # S5261; Sigma-Aldrich), 19.4 mg/l insulin (Cat. # A11382IJ; Thermo Fisher Scientific), 1383 mg/l NaHCO3 (Cat. # S5761; Sigma-Aldrich), 10.7 mg/l transferrin (Cat. # T0665; Sigma-Aldrich), 10 ml/l Glutamax (Cat. # 35050038; Thermo Fisher Scientific), 40µg/l FGF2-3 (Cat. # Qk053; Qkine) and 2 µg/l TGFβ1 (Cat. # 100-21; Peprotech). For time lapse experiments, E8 was prepared using DMEM/F12 without phenol red (Cat. # D6434; Sigma-Aldrich). Cells were fed daily and maintained at 37°C under a humidified atmosphere of 5% CO2 in air.   RNA was isolated, libraries constructed and sequenced by GENEWIZ (Azenta Life Sciences, UK). libraries were prepared for Illumina (New England Biolabs, Ipswich, USA) and the library preparations were sequenced on an Illumina Novaseq platform (Illumina, San Diego, USA) to generate 150 bp paired-end reads.