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| Status |
Public on Feb 02, 2017 |
| Title |
5-6hr_input |
| Sample type |
SRA |
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| Source name |
embryo
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| Organism |
Drosophila melanogaster |
| Characteristics |
strain: wild type
antibody: none
Stage: 5-6 hours after egg laying
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| Growth protocol |
For ChIP-seq experiments, wild type OreR embryos at various stages were collected. To deplete zygotic GRH, two mutants (grhIM and grhB37) were balanced over CyO, sChFP and embryos were scored for fluorescence. To deplete maternal GRH, germline clones were produced as described in Harrison et al., 2010 using the FLP-FRT system described in Chou and Perrimon, 1996
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP-seq: Embryos were collected, fixed in formaldehyde, chromatin was isolated, and the chromatin was immunoprecipitated as described in Li et al., 2008. Immunoprecipitation was done using two antibodies against Grainy head and an input sample was also collected for sequencing. RNA-seq: Embryos depleted of either maternal or zygotic GRH as well as sibling controls were collected and examined to determine the stage. 7 embryos were collected for each set and lysed in TRIzol reagent (Life Technologies) supplemented with 150 µg/ml glycogen. RNA was extracted and cDNA libraries were prepared using the Illumina Truseq RNA sample prep kit.
ChIP-seq libraries were prepared using standard Illumina protocols and RNA-seq libraries were prepared using Illumina Truseq RNA sample prep kit according to manufacterer's instructions
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Basecalls performed using CASAVA1.8.2
All analyses were done on the Galaxy online platform
Raw ChIP reads were trimmed for quality by sliding window with a minimum score of 28
ChIP reads were aligned to the dm3 genome assembly using Bowtie with maximum of 2 mismatches and unmapped reads were removed. Additional mapping was done using the dm6 genome and these re-mapped sequences are provided as bam files.
ChIP peaks were called with MACS with a p-value cutoff of 1e-6, MFOLD renrichment ratio of 5, Tag size of 80, and calculating the maximum lambda as local lambda at 1000, 2500, and 5000bp
Raw RNA-seq reads weere trimmed for quality by sliding window with a minimum score of 28
Reads were aligned to the dm3 geneome assembly using TopHat using default settings
Cufflinks was used to assemble transcripts and estimate FPKM for TopHat-aligned data
Cuffmerge was used to merge together resultant Cufflinks files (as GTF files)
Fold change between experimental conditions (with or without GRH) was calculated using CuffDiff, using the TopHat and Cuffmerge files produced in previous steps.
Genome_build: dm3 or dm6
Supplementary_files_format_and_content: BED: File produced by MACS peak calling software. File contains genomic regions corresponding to ChIP peaks (mapped to dm3)
Supplementary_files_format_and_content: WIG: File produced by MACS peak calling software. Contains profile data of genome from ChIP-seq experiment (mapped to dm3)
Supplementary_files_format_and_content: TABULAR: File produced by CuffDiff softare for assessing RNA-seq data. Fold change, p-value, and q-values are given for each gene or locus that describes the change in gene expression between experiment conditions.
Supplementary_files_format_and_content: BAM: File produced by Bowtie mapping software. File is mapped to "dm6" Drosophila genome
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| Submission date |
Jun 13, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Melissa M Harrison |
| E-mail(s) |
[email protected]
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| Organization name |
University of Wisconsin Madison
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| Department |
Biomolecular Chemistry
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| Lab |
1135 Biochemical Sciences Bldg
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| Street address |
420 Henry Mall
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| City |
Madison |
| State/province |
WI |
| ZIP/Postal code |
53706 |
| Country |
USA |
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| Platform ID |
GPL13304 |
| Series (1) |
| GSE83305 |
Stable Binding of the Conserved Transcription Factor Grainy Head to Its Target Genes Throughout Drosophila melanogaster Development |
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| Relations |
| BioSample |
SAMN05238503 |
| SRA |
SRX1840076 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2199206_5-6hr_INPUT_mapped_dm6.bam |
954.7 Mb |
(ftp)(http) |
BAM |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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