title: GSM8147869: Zebrafish Oocyte, Stage III, rep1; Danio rerio; RNA-Seq
library_construction_protocol: Isolated oocytes were snap-frozen and RNA was extracted with TRIzol (Invitrogen #15596026) following manufacturer's protocols. 0-, 2-, 4- and 6-hpf zebrafish embryo RNA was extracted with Direct-zol RNA MicroPrep kit (Zymo Research #R2062) following manufacturer's protocols. Ribo-dep Stranded RNA-Seq Zebrafish embryos (0-, 2-, 4-, and 6-hours post fertilization) total RNA sequencing libraries were generated from 500ng of high-quality total RNA, as assessed using Bioanalyzer (Agilent). Libraries were made according to the manufacturer's directions for the TruSeq Stranded Total RNA Library Prep Gold (Illumina #20020598), and TruSeq RNA Single Indexes Sets A and B (Illumina #20020492 and #20020493). Resulting short fragment libraries were checked for quality and quantity using the Bioanalyzer (Agilent Technologies) and Qubit Fluorometer (Life Technologies). Libraries were pooled, quantified, and sequenced as 75bp single reads on a high-output flow cell using the Illumina NextSeq500 instrument. Following sequencing, Illumina Primary Analysis (version RTA 2.11.3.0) and bcl-convert 3.10.5 were run to demultiplex reads for all libraries and generate FASTQ files. Zebrafish oocytes (stages I-IV) total RNA-Seq libraries were generated from 100 ng of high-quality total RNA, as assessed by a Bioanalyzer (Agilent Technologies). Libraries were prepared according to manufacturer's instructions using the TruSeq Stranded Total RNA Library Prep Gold (Illumina, Cat. No. 20020598), and TruSeq RNA Single Indexes; Sets A and B (Illumina Cat. No. 20020492 and 20020493). Resulting short-fragment libraries were checked for quality and quantity using a Bioanalyzer and a Qubit Fluorometer (Life Technologies). Libraries were normalized, pooled, and sequenced on a NextSeq 500 instrument (Illumina), as 75 bp single-end reads on a high-output flowcell using NextSeq Control Software 2.2.0.4. Following sequencing, Illumina Primary Analysis version NextSeq RTA 2.4.11 and Secondary Analysis version bcl2fastq2 v2.20 were run to demultiplex reads for all libraries and generate FASTQ files.