title: RNA-Seq of Danio rerio larvae: gnotobiotic 7 dpf
design: At 7 dpf, 4 zebrafish of condition were sampled and washed twice by 2 transfers to clean, sterile water in petri dishes. Each fish was then added to a sterile 2-mL microcentrifuge tube and euthanized with tricaine at 0.3 mg/mL. All liquid was removed from the tissue and the samples were immediately frozen at -80C until RNA extraction. RNA was extracted from whole single larvae using the Zymo Quick-RNA Micro Prep kit (Zymo Cat. R1050) with additional bead homogenization and proteinase K treatments. Purified RNA was quantified using the Qubit HS RNA fluorometer kit (ThermoFisher Q32851) and purity was assessed with the Nanodrop spectrophotometer (ThermoFisher). RNA sequencing libraries were prepared with Illumina TruSeq stranded with PolyA selection and sequenced using Illumina NovaSeq, 2x150bp paired-end (PE) reads. Target of 20 million PE reads per sample