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| Status |
Public on Dec 01, 2019 |
| Title |
SAFB-K9me3-HiChip-Rep2-plus |
| Sample type |
SRA |
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| Source name |
AML12 shSafb
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| Organism |
Mus musculus |
| Characteristics |
cell line: AML12
lentivirus: shSafb
antibody: H3K9me3
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| Treatment protocol |
The shRNA lentiviruses were prepared as previously described (Fu et al., 2015). Then AML12 cells were infected with 200 μl of virus concentrate in a well of six-well-plates. Medium was changed 24 hours after transfection, and then collect cells 5 days after transfection.
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| Growth protocol |
AML12 cells were cultured in DMEM/F12 supplemented with 10% fetal bovine serum, ITS Liquid Media Supplement, and 0.1μM dexamethasone at 37 °C and 5% CO2.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
The HiChIP of H3K9me3 was performed as described (Mumbach et al., 2016), started with 2.5 millions of AML12 cells for each sam- ple. Briefly, 2.5 millions of cells were digested by 100U of MboI overnight at 37 C. After biotin filling and DNA ligating, the nuclear were resuspended by sonication buffer (1%SDS, 10mM EDTA, 50mM Tris-HCl, 20mM NaBu, 1 3 Proteinase Inhibitor), and chromatin was sheared into 300-700bp using Bioruptor followed by incubating with antibody overnight at 4 C. 25 mL beads were washed once with cold 0.5% BSA, and added into the sheared chromatin-protein supernatant for 2 hours at 4 C. Washed with high salt wash buffer (50mM HEPEs, 500mM NaCl, 1mM EDTA, 0.1%DOC, 1%Triton X-100) three times; low wash buffer (10mM Tris-HCl, 250mM LiCl, 1mM EDTA, 0.5%NP40, 0.5%DOC) twice; TE once. Chromatin-protein complex were eluted for 15min at 65 C in 50 mL elution buffer (1% SDS in TE). After reverse-crosslinking, DNA was extracted using ultrapure phenol/chloroform. Chromatin fragments con- taining biotin were captured by MyOne Streptatvidin beads (ThermoFished 65601), and libraries were prepared on beads using Tn5 from DNA Library Prep Kit V2 (Vazyme TD503) for 30min at 37 C, followed by amplification.
Illumina sequencing library were prepared according to the standard Illumina protocol.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
HiSeq X Ten |
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| Data processing |
Library strategy: HiChIP
Hi-Chip paired-end reads were aligned to mm9 reference genome using the HiC-Pro pipeline (Servant et al., 2015) with default set- tings. We converted .all ValidPairs files to .hic files by the script hicpro2juicebox.sh from HiC-Pro utilities. To detect the differences between experiments, we normalized the interaction matrices for each chromosome separately according to the total contacts in each chromosome. This normalization led to the sum of contacts in each chromosome to 1e8.
Genome_build: mm9
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| Submission date |
Nov 27, 2019 |
| Last update date |
Dec 01, 2019 |
| Contact name |
Bo Wen |
| E-mail(s) |
bowen75@fudan.edu.cn
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| Organization name |
Fudan University
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| Street address |
130 Dongan Rd.
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| City |
ShangHai |
| ZIP/Postal code |
200032 |
| Country |
China |
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| Platform ID |
GPL21273 |
| Series (2) |
| GSE125037 |
The Nuclear Matrix Protein SAFB Cooperates with Major Satellite RNAs to Stabilize Heterochromatin Architecture Partially through Phase Separation |
| GSE141113 |
The nuclear matrix protein SAFB maintains heterochromatin architecture through RNA-dependent phase separation [HiChIP] |
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| Relations |
| BioSample |
SAMN13412358 |
| SRA |
SRX7228451 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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