title: RNA-Seq of Cryptotermes secundus, primary queen, > 4 years old, head and thorax tissue, Sample ID EE36-1c
design: Examination of gene/transcript expression patterns in primary queens of the termite Cryptotermes secundus (different ages examined, head and thorax). Individuals were placed on ice and the gut was removed and discarded. The head together with the thorax were used for RNA extraction. Samples were transferred into peqGOLD Tri Fast (PEQLAB) and homogenized in a Tissue Lyser II (QIAGEN). Chloroform was used for protein precipitation. From the aqueous phase, RNA was precipitated using Ambion isopropyl alcohol and then washed with 75% ethanol. Obtained pellets were solved in nuclease-free water. DNA was subsequently digested using the DNase I Amplification Grade kit (Sigma Aldrich, Cat. No. AMPD1). A RIN Analysis was performed measuring the RNA concentration with the Agilent RNA 6000 Nano Kit using an Agilent 2100 Bioanalyzer (Agilent Technologies) for quality control. Samples with total RNA were sent on dry ice to Beijing Genomics Institute (BGI) Tech Solutions (HONGKONG) Co. and then to the BGI-Shenzhen (PR China) for sequencing. The preparation of the cDNA libraries was performed by BGI according to their internal and proprietary standard operating procedure. The cDNA libraries were paired-end sequenced (not-strand specific) on Illumina HiSeq 4000 platforms (100 base pairs read length and about 4 Giga bases per sample). Index sequences from the machine reads were demultiplexed and a quality -check and filtering of raw reads was done using the package soapuke (-n 0.05 -l 20 -q 0.2 -p 1 -i -Q 2 -G --seqType 1 and -A 0.5, http://soap.genomics.org.cn/).Worker were flash frozen in liquid nitrogen after decapitation. Heads and abdomens were transferred and stored at -80C before dissection of brain and fat body tissue. Tissues dissected on ice in 1% PBS. The brain was dissected within less than five minutes, immediately homogenised in 50 L of Trizol and then stored at -80C until RNA extraction. Total RNA was extracted from each sample separately with the RNeasy mini kit (Qiagen). cDNA libraries were constructed with the transcriptome TruSeq RNA Library Prep Kit v2 (Illumina) and the SuperScript II Reverse Transcriptase (Thermo Fisher Scientific).