title: RNA-seq of tost-1(xf196 ts) mutant and wild-type embryos at 15°C and 25°C
design: RNA-seq of tost-1(xf196 ts) mutant and wild-type embryos at 15°C and 25°C
library_construction_protocol: Synchronized gravid adults were subjected to hypochlorite treatment to isolate embryos. Embryos were washed four times with cold M9 buffer and snap-frozen as \"worm balls\" in liquid nitrogen.  C. elegans strains were maintained at 20°C on NGM plates seeded with E. coli OP50. Animals were synchronized by hypochlorite treatment and hatched in M9 buffer for 16 hours. Synchronized L1 larvae were transferred to either 15°C or 25°C incubators and grown to gravid adult stage. Embryos were harvested by hypochlorite treatment when the first laid eggs were observed on plates, approximately 3-4 days post-L1 transfer depending on temperature. Frozen samples were ground with mortar and pestle, then mixed with five volumes of TRIzol™ LS Reagent (Invitrogen™). RNA was extracted using Direct-zol RNA Microprep kit (Zymo Research™) with additional TURBO DNase treatment (Invitrogen™) to remove genomic DNA. RNA was resuspended in nuclease-free water. NGS libraries were prepared using Illumina's Stranded mRNA Prep Ligation Kit with 1000 ng starting material and 10 amplification cycles. ERCC spike-ins (2 μl of 1:100 dilution, Ambion) were added to assess technical variability and optionally assist with library size normalization. Libraries were profiled using DNA 1000 chip on 2100 Bioanalyzer (Agilent) and quantified with Qubit dsDNA HS Assay Kit. 
identifiers.namespace: European Bioinformatics Institute
identifiers.id: E-MTAB-15730:wt_25_rep3_s