P-MTAB-25971 - P-MTAB-25971
Tissue samples were treated prior to the isolation of nucleic acids with a phenol-chloroform extraction protocol. First, the samples were homogenized in Lysing Matrix D Tubes containing 700 uL RLT-buffer (Rneasy Lysis Buffer, 2-Mercaptoethanol added) with the FastPrep-24 Instrument from Biomedicals (3000×g, 3 min). The supernatants were transferred and 700 uL phenol-chloroform-isoamyl alcohol were added. Tubes were mixed and centrifuged at 12000×g for 5 min and then 500 uL chloroform-isoamyl alcohol were added and incubated for 3 min at room temperature. After another centrifugation step, the top phase was used for the extraction of RNA. Total RNA was isolated according to the Rneasy protocol from Qiagen (Cat#74182). RNA concentrations were analyzed by NanoDrop ND-1000 UV-Vis Spectrophotometer at 260 nm (Thermo Scientific). RNA purity was assessed with the 260/280 value (pure samples show a range of 1.8 – 2.1) and samples were stored at -80C.