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| Status |
Public on Sep 02, 2022 |
| Title |
11382_10331_111584_HT5YVBGXC_GAGGGG_HH6_1 |
| Sample type |
SRA |
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| Source name |
TFAP2AE1 GFP+ Sorted
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| Organism |
Gallus gallus |
| Characteristics |
time: HH6
condition: NC
group: 1
cell type: TFAP2AE1 GFP+ Sorted
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| Treatment protocol |
37C ex-ovo incubation, whole-embryo electroporation of ptk-TFAP2AE1-GFP, Cranial dissections, dissociation using Accumax, FACS sorting for GFP+ neural crest cells, frozen in CryoStor media (Stem Cell Technologies #CS10), processed using OMNI-ATAC-Seq protocol
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Group #1 - Cranial dissection and dissociation using Accumax, Group #2 - Half-head cranial dissection and dissociation using Accumax, Group #3 - culture dissociation using Accutase and counting to normalize cell input.
Cells were processed using the OMNI-ATAC-Seq protocol. Frozen cells were quickly thawed at 37℃ and ATAC-RSB buffer was added to a total volume of 1.5mL. Samples were centrifuged at 500rcf for 5 minutes at 4℃ and the supernatant was removed. Next, cells were resuspended in 100µl of ATAC-RSB-LYSIS and kept on ice for 3-5 minutes, depending on input cell type. To stop lysis, 1mL of ATAC-RSB-WASH was added to each sample and they were again centrifuged for 5 minutes at 500rcf. The supernatant was removed, and cells were resuspended in 50µl of OMNI-ATAC Mix (~100nM concentration of Illumina TDE1 enzyme). Cells were then tagmented on a mixing (500rpm) thermoblock at 37℃ for one hour. Tagmented DNA was recovered using a Qiagen MinElute Kit (#28204), with 21µl of elution buffer warmed to 55℃. Library amplification PCR was performed with the NEBNext Ultra II Q5 2X Master Mix (NEB #M0544S) using Nextera primers for 13-15 cycles.
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| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
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| Data processing |
Demultiplexing, bcl2fastq
Trimming, CutAdapt v2.10, -a CTGTCTCTTATACACATCT -A AGATGTGTATAAGAGACAG --minimum-length=25 -j 0
Alignment, bowtie2, (Groups #1,#2 bowtie2 --local --very-sensitive-local --no-unal --no-mixed --no-discordant --threads 16 -x ~/genome/bowtie2-GRCg6a/GRCg6a -X 2000), (Group #3, -x~/genome/hg38/hg38_bt2/GRCh38)
Mark Duplicates, picard MarkDuplicates
Filter Duplicates, samtools v1.9, samtools view -@ 16 -F 1804 -f 2
Call peaks, MACS2, (Groups #1,#2 macs2 callpeak -t "$FILE" -n "$FILE" -f BAMPE -g 1218492533 -q 0.05 --call-summits --nomodel --shift 37 --ext 73 -B --SPMR)(Group #3 -g 2913022398)
Combine datasets/R analysis, DiffBind
Genome_build: for Gallus gallus, ENSEMBL galGal6 (GRCg6a)
Genome_build: for Homo sapiens, UCSC hg38
narrowPeak, peak file from MACS2
.RDS, R data file, specifically the S3 Object of class DBA from DiffBind. Contains peak by sample count matrix.
.txt, tab-delim text file, peak by sample matrix for each experiment.
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| Submission date |
Dec 23, 2020 |
| Last update date |
Sep 02, 2022 |
| Contact name |
Marcos Simoes-Costa |
| E-mail(s) |
simoescosta@cornell.edu
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| Organization name |
Cornell University
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| Department |
Molecular Biology and Genetics
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| Lab |
Simoes-Costa Lab
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| Street address |
526 Campus Rd
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| City |
Ithaca |
| State/province |
New York |
| ZIP/Postal code |
14853 |
| Country |
USA |
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| Platform ID |
GPL27748 |
| Series (2) |
| GSE163771 |
OCT4-SOX2 dimers reshape the epigenome to promote neural crest multipotency [ATAC-Seq] |
| GSE163961 |
OCT4-SOX2 dimers reshape the epigenome to promote neural crest multipotency |
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| Relations |
| BioSample |
SAMN17146523 |
| SRA |
SRX9724100 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4986891_11382_10331_111584_HT5YVBGXC_GAGGGG_HH6_1.narrowPeak.gz |
1.4 Mb |
(ftp)(http) |
NARROWPEAK |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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