GSM8697718: WT,rep4; Mus musculus; RNA-Seq
Aortic valve tissue samples were collected in an RNase-free setting from wildtype, Notch1+/-;Gata5+/- and Notch1+/;Gata5-/- mice at P10. Aortic valve tissues were harvested in Trizol, snap frozen and homogenized using TissueLyser II. Three aortic valves tissue were pooled to form one biological replicate for each genotype. Total RNA was isolated using Norgen Bioteck total RNA isolation kit, and DNase treatment was performed post RNA purification. Messenger RNA was purified from total RNA 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. Quantified libraries were pooled and sequenced on Illumina platforms, according to effective library concentration and data amount.