biostudies-arrayexpressUnknownTranscriptomicsGenomicsProteomicsCarolin Schmidtketranscription profiling by arrayMus musculusMus musculushttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-5706Obtain insight into lysosomal signaling pathwaysbiostudies-arrayexpressHybridization - For hybridisation, the Agilent Gene Expression Hybridisation Kit (Agilent Technologies) is used. 600 ng of each cRNA is hybridised on 8x60K microarrays at 65 °C for 17 h using Agilent's recommended hybridisation chamber and oven. Afterwards microarrays are washed once with the Agilent Gene Expression Wash Buffer 1 for one minute at ambient temperature followed by a second wash with preheated (37 °C) Gene Expression Wash Buffer 2 for one minute.Labeling - For labelling, the Low Input QuickAmp Labeling Kit (Agilent Technologies) is used to generate fluorescent cRNA. For 1st strand synthesis, a random primer / oligo-dT primer mixture is used. After 2nd strand synthesis, an in vitro transcription for synthesis of cRNA labelled with cyanine 3-CTP is performed.Growth Protocol - cerebellar cells were grown on 3.5 cm plates in DMEM supplemented with 10 % FCS, 1x GlutaMax, 1x PenStrep, 24 mM KCl at 33 °C, 5 % CO2Nucleic Acid Extraction - RNA was isolated using Qiagen RNeasy columns according to the manufacturer's instructionsSample Collection - cells were washed two times with PBS on ice and lysed directly on the plate in RNA lysis buffer from Qiagen according to the manufacturer's instructionsMIAME ScoreRaw DataOrganizationAssays and DataMAGE-TAB FilesArray DesignsCarolin SchmidtkefalseTranscriptional profiling of Cln3 mutant cerebellar cell line against wild-typeObtain insight into lysosomal signaling pathways2022-12-30T00:00:00Z2022-01-31T19:49:37.726Z2022-01-31T19:49:37.726ZE-MTAB-5706EFO_0002768EFO_0002944EFO_0003813EFO_0003789EFO_0005518EFO_0003815