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Sample GSM5621132 Query DataSets for GSM5621132
Status Public on Aug 04, 2022
Title HP1g_KCl_0min_exp2
Sample type SRA
 
Source name E15.5 C57BL/6 embryonic mouse cortices
Organism Mus musculus
Characteristics strain: C57BL/6
age: E15.5
cell type: primary corticol neurons
chip antibody used for chip: Millipore 05-690 (Lot#3224566)
Treatment protocol For ChIP-seq experiments, mouse cortical neurons were plated at an approximate density of 2 × 10^6 on 35-mm dishes. Neurons were plated in 2 mL neuronal medium. One ml of the medium was replaced with 1 ml fresh warm medium on Day3 and Day6. Prior to KCl depolarization, neurons were silenced with 1 μM tetrodotoxin on Day6 (TTX; Fisher). Neurons were subsequently stimulated on Day7 by adding warmed KCl depolarization buffer (170 mM KCl, 2 mM CaCl2, 1 mM MgCl2 and 10 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES)) directly to the neuronal culture to a final concentration of 31% in the neuronal culture medium within the culture plate or well for the indicated time.
Growth protocol Neurons were grown in neuronal medium consisting of Neurobasal medium containing B27 supplement (2%; Thermo Fisher), penicillin-streptomycin (50 g/mL) and Glutamax (1x; Thermo Fisher). Neurons were subsequently plated and placed in a cell culture incubator that maintained a temperature of 37 °C and a CO2 concentration of 5%. Ten minutes after plating neurons, the medium was completely aspirated from cells and replaced with fresh warm neuronal medium. Neurons were grown in vitro until Day7.
Extracted molecule genomic DNA
Extraction protocol Cells were cross-linked with 1% formaldehyde at room temperature for 10 min. And then the cross-linking was quenched with 0.125M glycine for 5 min. Chromatin was fragmented using a Bioruptor® Pico (Diagenode) for 10 min at high power, with an interval of 30 s between pulses to get around 200bp fragments and precleared using 20 μl Protein G Dynabeads (Life Technologies, Cat# 10009D). Subsequently, the soluble chromatin was incubated with 2-5 μg antibodies at 4°C overnight. Immunoprecipitated complexes were collected using 30 μl Protein G Dynabeads, which have been saturated with PBS/1% BSA over night at 4°C, per reaction. For all ChIPs, after de-crosslinking overnight at 65°C, final ChIP DNA was extracted and purified using QIAquick spin columns (QIAGEN).
The libraries were constructed following Illumina’s Chip-Seq Sample prep kit.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina NovaSeq 6000
 
Data processing Basecalls performed using CASAVA version 1.4
Reads were aligned to the mm10 genome assembly using Bowtie2.
Genome_build: mm10
Supplementary_files_format_and_content: bedGraph
 
Submission date Oct 11, 2021
Last update date Aug 04, 2022
Contact name Yuliang Tan
E-mail(s) yut020@ucsd.edu
Organization name University of California, San Diego
Department School of Medicine
Street address Room 345, CMM West, 9500 Gilman Drive, Mail Code 0648
City La Jolla
State/province California
ZIP/Postal code 92037-0648
Country USA
 
Platform ID GPL24247
Series (2)
GSE135808 Topoisomerase 1 Nicking-dependent HP1gamma Engagement Mediates Acute Activation of Phase-separated Enhancers
GSE185648 Ku70/HP1g are presented at the KCl induced acutely activated enhancers [ChIP-seq]
Relations
BioSample SAMN22213273
SRA SRX12569174

Supplementary file Size Download File type/resource
GSM5621132_HP1g_KCl_0min.bed.gz 591.4 Kb (ftp)(http) BED
GSM5621132_HP1g_KCl_0min_exp2.ucsc.bedGraph.gz 384.7 Mb (ftp)(http) BEDGRAPH
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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