title: GSM5099492: M157vmn; Coturnix japonica; RNA-Seq
library_construction_protocol: Brains were sliced in 200um-thick coronal sections in a 2800 Frigocut E cryostat (Reichert-Jung) and the sections kept on Superfrost Plus (Thermo Scientific,) microscope slides previously cooled to the temperature of the cryostat. Brain nuclei were punched using a specialized 1.25mm diameter brain punch apparatus (Leica Biosystems) using the previously observed neuroanatomical distribution of quail aromatase-positive cell populations as a guide. RNA was extracted using the RNeasy Mini Kit (Qiagen, Venlo, Netherlands), DNA digested on-column using the RNase-free DNase Set (Qiagen), RNA quantity measured on a NanoDrop 2000 spectrophotometer (Thermo Scientific), concentrated by vacuum centrifuge, resuspended in 10ul RNase-free water (Ambion), and quantified again on Nanodrop. RNA samples underwent ribosomal depletion using Ribozero (Illumina, San Diego, CA), cDNA synthesis, barcoding, pooling by brain region (3 pools, each with n=12 samples/pool), and each pool was divided and run across all 4 lanes of a NextSeq500 chip (Illumina) for a total of three chips at the GIGA-Genotranscriptomics platform (ULg, Liege Belgium). The cDNA were read in paired-end, 75nt read length format, and adapter and barcode sequences trimmed prior to assembly.