title: GSM4190540: 2534_E: Mouse 1 NI clone 1; Mus musculus; RNA-Seq
library_construction_protocol: Mechanically disrupt organoids and Matrigel with cold 1X PBS and transfer into a 15ml falcon  tube containing 10ml cold 1X PBS. Centrifuge at 300g for 5 minutes at 4°C. Remove supernatant  and add 1ml of Trizol to the pellet and transfer to a 1.5ml tube and keep overnight or longer at -80°C. Defreeze the samples at RT and add 200µl of chloroform. Mix the tubes for 15 seconds (do no vortex) and incubate at RT for 5 min. Centrifuge at 13000g for 15 minutes at 4°C. Transfer the top aqueous phase to a new 1.5ml tube (do not transfer anything from the interphase). Add 500µl of isopropanol, 120µl of 5M ammonium acetate, and 10µl of glycogen (5mg/ml). Vortex briefly and store for a minimum of 15 minutes at -20°C. Centrifuge at 13000g for 15 minutes at 4°C and discard the supernatant. Wash the pellet with 1ml 75% cold ethanol and vortex until you see the pellet floating. Centrifuge again at 13000g for 10 minutes at 4°C. Carefully discard the supernatant and air dry the pellet. Dissolve the pellet in 50µl of nuclease free H2O. Measure the concentration at the NanoDrop and store at -80°C. cDNA was generated from polyA enriched RNA.
attributes.tag: GEO Accession
attributes.value: GSM4190540