title: Transcriptome sequencing of Gryllus bimaculatus 3rd instar
design: ample preservation, RNA extraction, sequencing and assembly were conducted using methods previously described for spiders [1], with some modifications. Briefly, a whole body of the single specimen of each of the developmental stages of G. bimaculatus was snap-frozen with liquid nitrogen and stored at -80C, and RNA was extracted using TRIzol reagent (Thermo Scientific) after homogenization with a metal cone using the Multi Beads Shocker (Yasui Kikai). RNA was further purified using RNeasy Plus Mini Kit (Qiagen). Purified total RNA was quantified with a NanoDrop 2000 (Thermo Scientific) and a Qubit Broad Range (BR) RNA assay (Life Technologies). The integrity of RNA was estimated by electrophoresis using TapeStation 2200 with RNA Screen Tape (Agilent Technologies). The sequence library was prepared using NEB Next Ultra II RNA Library Prep Kit for Illumina (NEB) according to manufacturers protocol, and then was sequenced on a NextSeq 500 (Illumina) with 300 cycles of high-output mode as paired-end reads. Sequences were base called and demultiplexed, and adaptor sequences were removed with bcls2fastq v.2 software (Illumina).