Short read sequencing of the transcriptome associated with spleen tissue in Dissostichus eleginoides
RNA was extracted from individual tissues using the Direct-zol RNA Miniprep Plus kit (Zymo Research) according to the manufacturers instructions. Briefly, 50 mg of the toothfish tissue was ground to a powder with liquid nitrogen in a precooled mortar and pestle. and then lysed in 600 l of TRI Reagent. The lysed sample was transferred to a clean 1.5 microtube and an equal volume of absolute ethanol added. The sample was thoroughly mixed and 700 l transferred to the Zymo-Spin IIICG column and centrifuged at 13,000g for 30 sec. The flow through was discarded and the column treated with DNase 1 prior to washing with 400 l of Direct-zol RNA prewash, followed by a wash with 700 l RNA wash buffer. The RNA was then eluted in 100 l DNase/RNase free water and stored at -80C.  For short read sequencing, mRNA libraries were prepared using the TruSeq stranded mRNA sample preparation kit with TruSeq adapters (Illumina) to enable multiplexing of the libraries into two pools. These were sequenced on an Illumina NovaSeq to 100 bp in paired-end mode. Sequence library preparation and subsequent sequencing were conducted by the Exeter Sequencing Service (University of Exeter).