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| Status |
Public on Jan 20, 2022 |
| Title |
H3K4me3 ChIP in KO decidualized stromal cells |
| Sample type |
SRA |
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| Source name |
decidualized mouse stromal cells
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| Organism |
Mus musculus |
| Characteristics |
strain: C57
tissue: Mouse uteri on pregnant day 8
chip antibody: H3K4me3 Rabbit pAb, vendor:Abcam; catalog number: #ab8580; lot/batch number: GR134451-3
genotype/variation: KO
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| Growth protocol |
Mice were housed in the animal care facility of Xiamen University according to the guidelines for the care and use of laboratory animals. All experimental procedures were approved by the Animal Welfare Committee of Research Organization (X200811), Xiamen University.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Decidual tissue wetted with PBS quickly cut into small pieces with an iris scissor. The pieces should not be bigger than 0.5 cm3. Snipped tissue was fixed with 1% formaldehyde (CST) for 10 mins at RT (for H3K4me3 ChIP), or fixed with disuccinimidyl glutarate (DSG, Santa Cruz, 2 mM) for 30 mins at RT, washed twice with PBS and then double fixed with 1% formaldehyde for another 10 mins at RT (for Menin ChIP). Fixation was stopped by adding glycine (0.125 M). Tissue homogenate obtained by using Dounce Tissue Grinder was filtered through a 200 μm cell strainer to remove connective tissue. Fixed cells were lysed by Lysis Buffer 1 (50 mM HEPES pH 7.5, 1 mM EDTA, 140 mM NaCl, 0.5% NP-40, 10% glycerol, 0.25% Triton X-100) and Lysis Buffer 2 (10 mM Tris-HCl pH8.0, 1 mM EDTA, 0.5 mM EGTA, 200 mM NaCl) and Lysis Buffer 3 (10 mM Tris-HCl pH8.0, 1 mM EDTA, 0.5 mM EGTA, 100 mM NaCl, 0.1% Sodium Deoxycholate, 0.1% N-lauroylsarcosine). All lysis buffers should be supplemented with protease inhibitors cocktail (Roche) before use. Chromatin DNA was sheared to 300-500 bp average in size by using a BioRuptor sonicator (Diagenode). Solubilized chromatin was was centrifuged to remove debris and incubated overnight at 4°C with antibody against Menin (Bethyl, 10 μg) and H3K4me3 (Abcam, 1 μg) bound to 20 μl protein A magnetic beads (Invitrogen). After washing and elution, the protein-DNA complex was reversed by heating at 65°C overnight. Immunoprecipitated DNA was purified by using QIAquick spin columns (Qiagen). Immunoprecipitated and input DNA were quantified using Qubit 4.0 fluorometer. ChIP-seq libraries were prepared by using the KAPA DNA Hyper Prep Kit (KK8502) following the manufacturer’s instruction and sequenced with an Illumina Nova PE150. For ChIP-qPCR, chromatin was sheared by sonication until the average length of 500-1000 bp.
The ChIP libraries were prepared using KAPA HyperPrep Kits (Roche, 07962347001) and then run on the Illumina sequencer Hiseq-Xten PE150 using standard Illumina protocols
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
D8 decidua tissue
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| Data processing |
Illumina Casava1.7 software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm10 whole genome using STAR with default parameters
MACS2 was used to call the peaks of Menin and H3K4me3 binding using default parameters.
Genome_build: mm10
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| Submission date |
Dec 20, 2021 |
| Last update date |
Jan 20, 2022 |
| Contact name |
Wenbo Deng |
| E-mail(s) |
wbdeng@xmu.edu.cn
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| Phone |
+86 17850568375
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| Organization name |
Xiamen University
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| Department |
School of Medicine
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| Street address |
Xiang'an Road
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| City |
Xiamen |
| State/province |
Fujian |
| ZIP/Postal code |
361102 |
| Country |
China |
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| Platform ID |
GPL24247 |
| Series (1) |
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| Relations |
| BioSample |
SAMN24252915 |
| SRA |
SRX13460139 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5743704_H3K4me3-KO_bedgraph.tdf |
114.8 Mb |
(ftp)(http) |
TDF |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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