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Status |
Public on Nov 21, 2022 |
Title |
CUTRUN_State1_Klf4_BR1 |
Sample type |
SRA |
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Source name |
State1, Klf4
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Organism |
Mus musculus |
Characteristics |
cell line: V6.5 cell type: embryonic stem cell line cut&run antibody: Klf4 (Abcam ab12947) genotype: WT
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Growth protocol |
V6.5 mouse embryonic stem cells (mESC) were cultured on tissue culture-treated 10cm plates pre-coated with 0.2% gelatin in phosphate-buffered saline (PBS). ESC were cultured in Dulbecco’s Modified Eagle Medium (Fisher), plus 10% fetal bovine serum, 1x penicillin/streptomycin, 1x non-essential amino acids, 1x β-mercaptoethanol, and 1000 U ml-1 leukemia inhibitor factor. All cells were grown at 37C and 5% CO2 and passaged every two days to maintain 10-70% confluency.
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Extracted molecule |
genomic DNA |
Extraction protocol |
CUT&RUN was performed using CUTANA ChIC/CUT&Run Kit (Epicypher 14-1048) following manufacturer’s instruction in User Manual Version 2.1 or User Manual Version 3. Three technical replicates per antibody were performed. For sorted state populations, cells were harvested using versene, and 8x105 cells per replicate were isolated by flow cytometry. Harvested cells were permeabilized using 0.005% digitonin. Additionally, 1 ug of antibody per sample was added (Klf4 Abcam ab12947, Zfp281 Abcam ab10131), with Rabbit IgG antibody (CUTANA 13-0042) used as the negative control. E.coli spike-in DNA was also added to each sample.
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Library strategy |
OTHER |
Library source |
genomic |
Library selection |
other |
Instrument model |
Illumina NextSeq 500 |
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Description |
CUT&RUN for Klf4 in State 1
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Data processing |
CUT&RUN reads were trimmed using cutadapt (v1.16) (-m 10 -q 10). Paired-end reads were aligned to the mm10 reference genome and the E.coli MG1655 using bowtie2 with default parameters. The normalization factor for each replicate was calculated as 1/((unique E.coli reads⁄unique mouse reads)× 100%). Normalized bigWig files were generated for each replicate using the bamCoverage function in the deeptools package by inputting the normalization factor as the --scaleFactor parameter. Replicates were further normalized to IgG negative control using the bigwigCompare function with the parameter --operation log. Assembly: mm10 Supplementary files format and content: CUT&RUN: bigWig format Library strategy: CUT&RUN
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Submission date |
Sep 12, 2022 |
Last update date |
Nov 29, 2022 |
Contact name |
Sofia Hu |
E-mail(s) |
sofiahu@mit.edu, xsu@cpdr.org
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Organization name |
MIT
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Street address |
500 Main Street
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City |
Cambridge |
State/province |
MA |
ZIP/Postal code |
02139 |
Country |
USA |
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Platform ID |
GPL19057 |
Series (1) |
GSE169044 |
Transcription factor antagonism regulates heterogeneity in embryonic stem cell states |
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Relations |
BioSample |
SAMN30807455 |
SRA |
SRX17533786 |
Supplementary file |
Size |
Download |
File type/resource |
GSM6573865_CUTRUN_State1_Klf4_BR1.bigWig |
216.4 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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