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Sample GSM5623638 Query DataSets for GSM5623638
Status Public on Sep 29, 2022
Title ATAC-seq__limb_E115_wt_mouse_Rep-2
Sample type SRA
 
Source name forelimb
Organism Mus musculus
Characteristics Stage: E115
strain: C57Bl.6/J
genotype: wt
Treatment protocol ATAC-seq was performed as described previously (Buenrostro et al., 2015). Briefly, 1x105 isolated limb cells were employed per biological replicate. Cells were washed in cold PBS, lysed in fresh lysis buffer (10mM TrisCl pH7.4, 10mM NaCl, 3mM MgCl2, 0.1% (v/v) Igepal CA-630) for 2 min on ice, and finally pelleted for 10 min at 500 x g and 4°C. Following supernatant aspiration, nuclei-containing pellets were subjected to transposition using Tn5 Transposase for 30 min at 37° C.
Growth protocol Chicken, opossum and mouse limb buds were microdissected from embryos in cold PBS. Isolated limbs were then trypsinised 5 minutes at 37°C with continuous agitation with a P1000 pipette until no visible clumps remained. Limb cell suspensions were then passed through a 40 µm filter, centrifuged at 250 g for 5 min. Supernatants were then removed from isolated ESCs or limb cells which could then be used for downstream applications.
Extracted molecule genomic DNA
Extraction protocol Resulting DNA was then purified using MinElute Reaction Clean up kit, eluted in 11 µl of elution buffer and stored in -20° C, if not immediately processed further.
Barcoded adapters were added to the transposed fragments by PCR. To avoid saturation in our PCR, we initially performed 5 cycles and extracted a 5 µl aliquot for qPCR to identify the number of cycles required without overamplification. Nextera qPCR primers were used for the amplification. The remaining 45 µl of the PCR reaction were then amplified for the desired number of cycles which never exceeded 12. Finally, samples were purified on AMPure XP beads and eluted in 20 µl. Concentration was measured with Qubit and the quality of the samples was estimated by Bioanalyzer analysis. ATAC-seq samples were sequenced yielding for 50 million 75 bp paired-end reads and each experiment was performed in duplicate.
 
Library strategy ATAC-seq
Library source genomic
Library selection other
Instrument model Illumina HiSeq 2500
 
Data processing Raw sequencing fastq files were processed using cutadapt (Martin, 2011) for adapter trimming, Bowtie2 (Langmead and Salzberg, 2012) for mapping, SAMtools (Li et al., 2009) for filtering, sorting and removing duplicates, and deepTools (Ramirez et al., 2016) for generating coverage tracks.
Genome_build: mm10 galgal6 and mondom5
Supplementary_files_format_and_content: bigwig files
 
Submission date Oct 12, 2021
Last update date Oct 01, 2022
Contact name Michael I Robson
E-mail(s) robson@molgen.mpg.de
Organization name Max Planck Institute for Molecular Genetics
Street address Max Planck Institut für molekulare Genet, Ihnestrasse 63-73
City Berlin
ZIP/Postal code 14195
Country Germany
 
Platform ID GPL17021
Series (2)
GSE185774 Promoter repression and 3D-restructuring resolves divergent developmental gene expression in TADs [ATAC-Seq]
GSE185775 Repression and 3D-restructuring resolves regulatory conflicts in evolutionarily rearranged genomes
Relations
BioSample SAMN22232714
SRA SRX12581259

Supplementary file Size Download File type/resource
GSM5623638_ATAC-seq_forelimb_E115_mm10_wt_Rep2_L9152.cpm.bw 477.6 Mb (ftp)(http) BW
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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