|
Status |
Public on Nov 28, 2019 |
Title |
JONAS_2096: Non-transgenic IgG control sample |
Sample type |
SRA |
|
|
Source name |
Three day old etiolated seedlings
|
Organism |
Arabidopsis thaliana |
Characteristics |
tissue: seedlings treatment: Air 2h chip antibody: Goat IgG whole molecule epitope fusion tag: None
|
Treatment protocol |
Not treated
|
Growth protocol |
Seeds were sterilised using bleach then plated on 1.2% agar Linsmaier and Skoog medium plates. Stratification was at 4 celsius in the dark for three days. Plates were exposed to the light at room temperature for 2 h to induce germinaiton, then returned to the dark at 22 celsius. Plates were incubated for three days, after which treatments were conducted.
|
Extracted molecule |
genomic DNA |
Extraction protocol |
Conducted as described in O'Malley et al. 2016 (doi: 10.1016/j.cell.2016.04.038) and Chang et al. 2013 (10.7554/eLife.00675). Harvested seedlings were crosslinked with 1% formaldehyde under vacuum for 20 minutes then snap-frozen in liquid nitrogen. Nuclei were extracted from frozen, ground seedlings according to Qiao et al. 2016 (10.1126/science.1225974) then sonicated according to Chang et al .2016 using a Diagenode bioruptor to yield an average DNA fragment size of approximately 150-300 bases. Transcription-factor DNA complexes were isolated by immunoprecipitation with polyclonal anti-GFP raised in goat from David Dreschel (Max Planck Institute for Molecular Cell Biology and Genetics, Dresden, Germany) or, for non-transgenic controls, IgG from goat (supplied by Covance). These complexes were then retrieved using Protein G Dynabeads (Life Technologies). Conducted as described in Chang et al. 2013 (10.7554/eLife.00675) with one modification. Size selection was performed on SPRI beads, instead of on gels, with upper and lower cut-offs to yield a fragment size-range of 150-500 nucleotides. Sequencing was conducted on the Illumina HiSeq 2500 platform.
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|
|
Library strategy |
ChIP-Seq |
Library source |
genomic |
Library selection |
ChIP |
Instrument model |
Illumina HiSeq 2500 |
|
|
Data processing |
Base-calling was performed by Illumina Real-Time Analysis software v1.17 Reads were aligned by bowtie2 v2.0.5 using only the first 65 bases against the Arabidopsis thaliana TAIR10 reference genome. Shiftsize was calculated using phantompeakqualtools and default parameters Peaks called with MACS2 version 2.0.10 with parameters "callpeak -c --format=SAM --gsize=1.19e8 -p 99e-2 --nomodel --shiftsize=[from phantompeakqualtools] --down-sample --call-summits". A concatenated file of all non-transgenic control IP reads was used as the control in peak calling Genome_build: TAIR10 Supplementary_files_format_and_content: Summit region output created by MACS2 in bed format.
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|
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Submission date |
Aug 10, 2018 |
Last update date |
Nov 28, 2019 |
Contact name |
Mathew G Lewsey |
E-mail(s) |
m.lewsey@latrobe.edu.au
|
Organization name |
La Trobe University
|
Department |
Animal, Plant and Soil Sciences
|
Street address |
AgriBio, 5 Ring Rd
|
City |
Bundoora |
State/province |
VIC |
ZIP/Postal code |
3086 |
Country |
Australia |
|
|
Platform ID |
GPL17639 |
Series (1) |
GSE114430 |
ChIP-Seq of Arabidopsis Transcription Factors MYC2 and MYC3 |
|
Relations |
BioSample |
SAMN09791880 |
SRA |
SRX4528706 |