Synchronized day 1 adult wild-type (N2) worms were either collected immediately (0-h 20 °C control), or shifted to 28 °C HS for 1, 24, or 48 h before collection. RNA was collected as described previously [63]. Library preparation, sequencing, and initial data analyses were performed by Novogene. Total RNA was poly-A selected and then subjected to 150-bp paired-end Illumina HiSeq sequencing using three biological replicates for each condition. The raw reads were filtered to remove reads with adaptor contamination, reads consisting of > 10% uncertain nucleotides, and reads where > 50% of the bases were low quality (base quality < 20). The remaining clean reads (~ 97% for each sample) were mapped to the WBcel235 C. elegans reference genome. Approximately 92–93% of the clean reads were mapped per sample. Genes that had no mapped reads for control or HS conditions were excluded from the differential expression analysis. Differentially expressed genes were identified using DESeq (v.1.18.0) by normalizing reads based on the negative binomial distribution method and comparing each HS timepoint to the 0-h control.

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