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| Status |
Public on Apr 13, 2010 |
| Title |
ap2_ctrl_rep1 |
| Sample type |
SRA |
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| Source name |
influorescence
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| Organism |
Arabidopsis thaliana |
| Characteristics |
strain: Col-0 SALK_071140
cell type: influorescence
chip antibody: AP2 custom antibody raised against the C-terminus of AP2 amino acids 289-432 (right after the second AP2 DNA binding domain until the stop codon) produced in the lab of Prof. Xuemei Chen, UC Riverside.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
The entire experiment from harvest through deep sequencing was repeated twice to produce independent biological replicates. ChIP-seq was performed with an antibody raised against the C-terminus of AP2 amino acids 289-432 (right after the second AP2 DNA binding domain until the stop codon). Col-0 plants were used to determine binding sites in the fully native context and ap2-12 (SALK_071140) plants were used as negative controls for any non-specific background antibody binding and pulldown. To maximize AP2 levels, we harvested clusters of young influorescences at the beginning of bolting, but before flowers opened, when AP2 expression is high. Briefly, clusters of just-bolting inflorescences were harvested and fixed as described previously {Gómez-Mena et al., 2005, Development, 132, 429-38}. Frozen tissue was ground, filtered three times through Miracloth (Calibrochem), and washed as described previously thorough buffers M1, M2, and M3 {Gómez-Mena et al., 2005, Development, 132, 429-38}. Nuclear pellets were resuspended in sonic buffer as described (1 mM PEFA BLOC SC (Roche Diagnostics) was substituted for PMSF), split into technical duplicate samples, and sonicated with a Branson sonifier at continuous pulse (output level 3) for 8 rounds of 2 x 6 seconds and allowed to cool on ice between rounds. IP reactions were performed by incubating chromatin with 2.5 microliters anti-rabbit AP2 antiserum overnight at 4C as described {Gómez-Mena et al., 2005, Development, 132, 429-38}. The immunoprotein–chromatin complexes were captured by incubating with protein A-agarose beads (Santa Cruz Biotechnology), followed by consecutive washes in IP buffer and then elution as described {Gómez-Mena et al., 2005, Development, 132, 429-38}. Immunoprotein-DNA was then incubated consecutively in RNase A/T1 mix (Fermentas) and Proteinase K (Roche Diagnostics) as described after which DNA was purified using Minelute columns (Qiagen) {Gómez-Mena et al., 2005, Development, 132, 429-38}. ChIP samples we tested for enrichment by QPCR and then deep sequencing libraries were produced by standard Illumina protocols.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina Genome Analyzer II |
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| Data processing |
Standard Illumina base calling software was used to base call the 42 nucleotide sequence reads. We used SHORE (http://1001genomes.org/downloads/shore.html) for read mapping and coverage analysis. The raw reads were pruned of low quality bases at their 3’ end before mapping them to the TAIR9 genome, allowing for up to four mismatching nucleotides and no gaps.
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| Submission date |
Apr 12, 2010 |
| Last update date |
Sep 04, 2019 |
| Contact name |
Felix Ott |
| Organization name |
Max Planck Institute for Developmental Biology
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| Street address |
Spemannstr. 35-39
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| City |
Tuebingen |
| ZIP/Postal code |
72076 |
| Country |
Germany |
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| Platform ID |
GPL9302 |
| Series (1) |
| GSE21301 |
Orchestration of development by the bifunctional transcription factor, APETALA2 |
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| Relations |
| Reanalyzed by |
GSE136843 |
| SRA |
SRX019320 |
| BioSample |
SAMN00011709 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM532327_AP2-BioRep1-control.bed.gz |
600.8 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
| Raw data are available in SRA |
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