|
| Status |
Public on Jan 16, 2019 |
| Title |
Etv6_ChIP1 |
| Sample type |
SRA |
| |
|
| Source name |
Somitic cells
|
| Organism |
Xenopus laevis |
| Characteristics |
tissue: somites
Stage: stage 22
strain: J-strain
chip antibody: Etv6 (custom made by NovoPro Bioscience Inc, Shanghai, China)
|
| Treatment protocol |
The somites of stage 22 Xenopus embryos were manually dissected in 0.35x MMR using forceps. Crosslinker EGS was added into cell homogenates (EGS final concentration: 2 mM) before fixation (10 minutes) . Sonication was performed using Covaris S220 (Duty factor: 10%; cycle per burst: 200; intensity: 5; duration time: 300 seconds; cycle: 3).
|
| Growth protocol |
Xenopus embryos were obtained by artificial fertilisation. They were cultured at 19 degree in 0.1x MMR (1xMarc’s Modified Ringer: 100 mM NaCl, 2 mM KCl, 1 mM MgCl2, 2 mM CaCl2, 5 mM HEPES, pH 7.5) after being dejellied in 2% cysteine (PH 7.8-8.0).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Etv6 immunoprecipitated DNA was eluted from beads in a thermomixer at 65 degres for 15 min and the eluates and reserved inputs were reverse cross-linked overnight at 65C. The samples were then treated with RNaseA, proteinase K and phenol/chloroform purified, and quantified using Qubit Fluorometer
Sequencing libraries were constructed with 1 ng of DNA using NEBNext Ultra DNA Library Prep Kit (NEB#E7370) according to manufacturer’s protocol
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Base calling performed on illumina 1.9 platform
Raw sequence reads were checked for base quality, trimmed and filtered to exclude adapters using Trimmomatic (Version 0.32)
Filtered read were aligned to the X. Laevis V9.1 with BWA version 0.7.12
Peaks were analyzed using MACS2 peak calling software at default thresholds, with the input samples as control for each replicate.
Peaks consistent between replicates were identified using DiffBind (Bioconductor package)
Peaks located in TSS regions were identified using Homer software and used to perform Transcription factors binding site analysis with Homer software (findMotifsGenome.pl)
Genome_build: Xenopus Laevis V9.1
Supplementary_files_format_and_content: xlsx file contains Etv6 ChIP consistent peaks in triple biological replicates; bam files include RPKM value of each sample.
|
| |
|
| Submission date |
Jun 01, 2018 |
| Last update date |
Jan 16, 2019 |
| Contact name |
Lei Li |
| E-mail(s) |
lei.li@imm.ox.ac.uk
|
| Organization name |
WIMM
|
| Department |
Radcliffe Department of Medicine
|
| Street address |
University of Oxford
|
| City |
Oxford |
| ZIP/Postal code |
OX3 9DS |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL21248 |
| Series (2) |
| GSE115224 |
Etv6-dependent positive and negative gene regulatory network controls vegfa expression in vivo [ChIP-seq] |
| GSE115225 |
Etv6-dependent positive and negative gene regulatory network controls vegfa expression in vivo |
|
| Relations |
| BioSample |
SAMN09294257 |
| SRA |
SRX4153521 |
