GSM8486522: Control_rep3; Malassezia restricta; RNA-Seq
Samples were concentrated using centrifugation (10,000 xg for 10 minutes) and added with 300ul of TRIzol™(Invitrogen) with rocking for 5 minutes, resuspended in trizol and stored at -80c. For RNA extraction, thawed samples were lysed by bead beating (Lysing Matrix E, 2 mL, MP Biomedicals) with three repeated cycles at 6,.5 m/s for 45 seconds. Subsequently, 200ul of Chloroform (Merck) was added per 1ml of trizol followed by centrifugation at 12,000 xg for 10mins. RNA was extracted using chloroform: isoamyl alcohol (24:1) (Merck), followed by precipitation by isopropanol (Merck). The final RNA pellet was washed with 70% ethanol followed by air drying for 2 hrs followed by dissolving in nuclease free water (Invitrogen). RNA libraries were prepared for sequencing using standard Illumina protocols