Cryoprotective Dehydration Experiment 1
Total RNA was extracted from larvae using Trizol reagent (Life Technologies, Grand Island, NY) according to the manufacturer’s protocol. RNA quantity and purity was assessed on a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA), and integrity was measured on an Agilent Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA). To generate RNA-seq libraries, we used the Illumina TruSeq RNA Sample Preparation kit (Illumina, San Diego, CA) according to the manufacturer’s protocol. In short, mRNA was purified from 2 µg total RNA from each sample, fragmented, and converted to double-stranded cDNA. Sequencing barcodes were ligated to the cDNA fragments, and the resulting fragments were amplified using PCR. Libraries were validated on an Agilent Bioanalyzer 2100 to ensure the libraries had the expected fragment size of ~300 bp.