Experiment information
Accession CRX048019
Organism Danio rerio
Title RIP-BS_0hpf_Ybx1-RIP-BisSeq
BioProject PRJCA001013
BioSample SAMC067647
Platform Illumina HiSeq 2500
Library name Construction protocol Strategy Source Selection Layout
Purified mRNA was used for RNA-BisSeq library construction. Around 200 ng mRNA premixed with the in vitro transcribed Dhfr mRNA at a ratio of 300:1 (Dhfr mRNA serves as methylation conversion control) was fragmented to ~100-nt fragments for 1 min at 90°C in 10× RNA Fragmentation Reagent (Ambion), then stopped by 10× RNA stop solution (Ambion), and precipitated with 100% ethanol. The RNA pellet was resuspended in 100 µl bisulfite solution (pH 5.1), which is a 100:1 mixture of 40% sodium bisulfite (Sigma) and 600 µM hydroquinone (Sigma) and subjected to heat incubation at 75°C for 4.5 h. The reaction mixture was desalted by passing through Nanosep with 3K Omega 500/pk columns (PALL Corporation) with centrifugation. After washed with nuclease-free water and centrifuged for five times, the RNA was finally disolved in 75 ?l nuclease-free water and then desulfonated by incubation with an equal volume of 1 M Tris-HCl (pH 9.0) at 75 °C for 1 h. After ethanol precipitation, the RNA was resuspended in 11 µl of RNase-free water and subjected to library construction. Reverse transcription was carried out with superscript II Reverse Transcriptase (Invitrogen) and ACT random hexamers. The following procedures were performed with the KAPA Stranded mRNA-Seq Kit (KAPA) according to the manufacturer’s instructions.Libraries were sequenced using HiSeq2500 (Illumina) in paired-read mode, creating reads with a length of 150 bp. Bisulfite-Seq TRANSCRIPTOMIC PCR PAIRED
Processing Planned read length (bp) for mate 1: 150
Planned read length (bp) for mate 2: 150
Insert size (bp): 240
Release date2019-06-12
Run Accession Release date Run Data File Information
File nameFile size (MB)
CRR053073 2019-06-12 RIP-BS_0hpf_R1.fastq.gz
Submittersun baofa (sunbf@big.ac.cn)
OrganizationsBeijing Institute of Genomics, Chinese Academy of Sciences
Date submitted2019-04-12