GSM2570485: Endothelial_GFP-_rep2; Mus musculus; RNA-Seq
Cells were sorted into TRIzol-LS (Invitrogen) and pooled into three independent biological samples. The cells were lysed in TRIzol-LS and the aqueous phase was collected after adding chloroform. The aqueous phase was homogenized by 70% ethanol and RNA isolated using Qiagen RNeasy micro kit. Isolated RNA was treated with TURBO DNA-free DNA Removal Kit to remove contaminating DNA. mRNA was isolated from total RNA using NEBNext Poly(A) mRNA Magnetic Isolation Module (E7490, NEB), fragmented to average size of 300 nt, reverse transcribed to generate double-stranded cDNA, and followed by NEBNext Ultra RNA Library Prep Kit for Illumina (E7530, NEB) according to manufacturer's instructions including the optional double size selection procedure using Agencourt AMPure XP beads. Prepared libraries were quantified with dsDNA HS Kit (Q32851) for Qubit and the size distribution assessed using High Sensitivity DNA Kit (5067-462) Bioanalyzer.