title: Transcriptome profiling in WT and hTDP-43 worms at temperatures 20C and 25C
design: Transcriptome profiling in WT and hTDP-43 worms at temperatures 20C and 25C
library_construction_protocol: Standard conditions were used for C. elegans propagation at 20 oC (Brenner, 1974). Animals were synchronized by hypochlorite bleaching, hatched overnight in M9 buffer at 20 °C and subsequently cultured on NGM agar plates seeded with OP50. Worms were grown from L1 until adulthood D1 at either 20 °C or 25 °C. The following genotypes were used: OW1601: dvIs62[snb-1p::hTDP-43/3'long UTR + mtl-2p::GFP]X and OW1603: dvIs15[(pPD30.38)unc-54 (vector) + (pCL26)mtl-2p::GFP]. OW1601 is CL6049 and OW1603 is CL2122 but then backcrossed an additional 6 times. CL2122 and CL6049 were gifts from Chris Link Total RNA from Day 1 adult C. elegans was isolated using TRizol (Invitrogen; #15596018) according manufacturer's protocol. The RNA concentration was assessed with a NanoDrop 2000 spectrophotometer (Thermo Scientific) and to assess the RNA quality, a 2100 BioAnalyzer (Agilent) was used to determine the RNA integrity score (RIN score) of all samples (Agilent; RNA 6000 Pico kit; #5067-15130) Next, rRNA was depleted from all samples with the NEXTflex RiboNaut rRNA depletion kit (PerkinElmer; #Nova-512961) starting with 800ng of total RNA. The cDNA library was made with the NEXTflex Rapid Directional qRNA-seq kit- Set B (PerkinElmer; #Nova-5130-04D).