GSM4676181: NPN-4; Equus caballus; RNA-Seq
Total RNA was isolated from all CA samples using RNeasy Mini Kit (#74104; Qiagen), and DNA digestion was performed on-column using RNase-free DNase I (#79254: Qiagen), followed by cleanup procedures. All procedures were performed according to the manufacturer's instructions. After extraction, RNA concentration and quality were analyzed using Nanodrop 2000 spectrophotometer (#ND-2000; Thermo Fisher Scientific) and Agilent 2100 Bioanalyzer® (Agilent, Santa Clara, CA, USA). All samples had a 260/280 ratio >2.0 and RNA integrity number (RIN) > 8.0. A TruSeq Stranded mRNA library prep kit (Illumina, San Diego, CA) was used to prepare libraries for mRNA sequencing. Libraries were loaded onto an Agilent DNA 1000 chip and validated on an Agilent 2100 Bioanalyzer (Agilent). Quantitation was performed with the Illumina Library Quantification Kit, ABI Prism qPCR Mix from Kapa Biosystems. Libraries were run on a NextSeq 500 v2 (Illumina) 300cycles High Output kit in a 2x150 base pairs with paired-end reads.