|
| Status |
Public on Jun 15, 2015 |
| Title |
WK33-HA mock 14h input rep2 |
| Sample type |
SRA |
| |
|
| Source name |
4 week old mature leaves
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
tissue: leaves
ecotype: Col-0
genotype: WRKY33-HA
chip antibody: -
treatment: 14 hour mock treated
|
| Treatment protocol |
For inoculation of Arabidopsis plants the spores were diluted in Vogel buffer. 2.5x10 to the power of 5 spores mL-1 were used for spray inoculations of 4-week old intact plants. For mock treatment, Vogel buffer was used. Plants were kept prior to and during infection under sealed hoods at high humidity.
|
| Growth protocol |
Arabidopsis Col-0 plants were grown for 4 weeks under short-day conditions in closed cabinets (Schneijder chambers: 16 h light/ 8 h dark cycle at 22-24°C, 60% relative humidity) on 42mm Jiffy-7 pots (Jiffy) to prevent contaminations from garden soil. Before sewing, the Jiffy pot peat pellets were re-hydrated in water containing 0.1% liquid fertilizer Wuxal (Manna). B. cinerea strain 2100 was cultivated on potato dextrose plates at 22°C for 10 days. Spores were collected, washed, and frozen at -80C in 0.8% NaCl at a concentration of 10 to the power of 7 spores mL-1.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Lysates were clarified from sonicated nuclei and WRKY33-DNA complexes were isolated with an HA antibody. ChIP DNA was purified using a QIA quick PCR Purification kit (Qiagen) and subjected to a linear DNA amplification (LinDA) protocol (Shankaranarayanan et al., 2011) which included two rounds of ‘in vitro transcription’ by T7 RNA polymerase.
The resulting LinDA DNA was used to generate sequencing libraries bearing barcodes using a NEBNext ChIP-Seq Library Pre Reagent Set for Illumina kit (New England Biolabs).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Negative control for peak calling only (for Sample 2)
|
| Data processing |
Illumina Casava1.8 software used for basecalling.
Sequencing read quality control was performed using FastQC version 0.11.2
Trimming and filtering of LinDA adapters and low quality sequences was performed using 1) cutadapt v1.2.1 with options –e 0.2, -n 2 and –m 36 and 2) PRINSEQ lite v0.20.2 with options –trim_qual_right/left 20, trim_tail_right/left 3 –min_len 36, -min_qual_mean 25
Sequenced reads were mapped to the Arabidopsis genome (TAIR10) using Bowtie v0.12.7 with options –best –m 1
Peak calling was performed for each replicate using the program QuEST v2.4 in transcription factor mode (option “2”) with permissive parameter settings (option “3”) and suppling the corresponding input or mock-treated sample as negative controls in peak calling
For samples where several negative controls were available, only peaks that were identified against all used negative controls (high confidence peaks) were retained for further analysis
Annotation of peak locations with respect to annotated gene features in TAIR10 was performed using the annotatePeaks.pl function from the Homer suite
Genome_build: TAIR10
Supplementary_files_format_and_content: tab-delimited text files with peak coordinates, enrichment scores, and annnotation information
|
| |
|
| Submission date |
Feb 25, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Barbara Kracher |
| E-mail(s) |
[email protected]
|
| Organization name |
Max Planck Institute for Plant Breeding Research
|
| Department |
Plant-Microbe Interactions
|
| Street address |
Carl-von-Linné-Weg 10
|
| City |
Cologne |
| ZIP/Postal code |
50829 |
| Country |
Germany |
| |
|
| Platform ID |
GPL17639 |
| Series (2) |
| GSE66289 |
WRKY33 binding sites in Arabidopsis upon Botrytis cinerea 2100 inoculation |
| GSE66300 |
Analysis of WRKY33 binding sites and WRKY33-dependent gene expression in Arabidopsis thaliana upon Botrytis cinerea 2100 inoculation |
|
| Relations |
| BioSample |
SAMN03375096 |
| SRA |
SRX890386 |
