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| Status |
Public on Feb 29, 2024 |
| Title |
Uterus,SMAD1-KI,PR,rep2 |
| Sample type |
SRA |
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| Source name |
Uterus
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| Organism |
Mus musculus |
| Characteristics |
tissue: Uterus
genotype: SMAD1-HA/HA
treatment: 4.5 dpc
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Uteri were harvested from two pregnant mice at 4.5 days post coitus and washed with cold swelling buffer (10 mM Tris-HCl pH 7.5, 2 mM MgCl2, 3 mM CaCl2, 1X Protease Inhibitor Cocktail (PIC, Roche, 11836170001)) immediately after collection. Then tissue was cut into small pieces (~2-3mm) using scissors, while submerged in cold swelling buffer. Nuclear extract was prepared by dounce homogenization in cold swelling buffer (using a size 7 dounce) and filtered using the cell strainer (100 μm, BD Biosciences). Lysate was centrifuged at 400 g for 10 min, then resuspended in lysis buffer (swelling buffer with 10% glycerol and 1% CA-630, 1X PIC) using an end-cut or wide-bore tips and incubated on ice for 5 min. Nuclei were washed twice with lysis buffer and resuspended in lysis buffer and incubated with activated concanavalin A beads for 10 min at room temperature. Bead-nuclei complexes were then resuspended in antibody buffer containing 0.01% digitonin, 1ug of each antibody was added to individual aliquots, and tubes were rotated at 4°C overnight. The following day, targeted chromatin digestion and release were performed with 2.5 ul of Cutana pA/G-MNase and 1 ul of 100 mM CaCl2. Genomic DNA with 0.5ng of Spike-in DNA together was purified with phenol-chloroform and ethanol precipitatation method.
Sequencing libraries were prepared using NEBNext Ultra II DNA Library Prep Kit (NEB E7645) following manufacture's protocol.
CUT&RUN-Seq
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Basecalls were performed using bcl2fastq v2.20 for NextSeq500 output.
Raw sequencing reads were aligned to the mm10 reference genome using bowtie2/2.2.7 in paired-end mode with parameters --end-to-end --very-sensitive --no-mixed --no-discordant --phred33 -I 10 -X 700
Raw sequencing reads for Spike-in were aligned to the E. coli genome U00096.3 using bowtie2/2.2.7 in paired-end mode with parameters --end-to-end --very-sensitive --no-mixed --no-discordant --phred33 -I 10 -X 700
Duplicated reads were removed, and only uniquely mapped reads were kept.
Spike-in normalization was achieved through multiply primary genome coverage by scale factor (100000 / fragments mapped to E. coli genome).Then normalized alignments were converted into fragments using bedtools bamtobed(v2.29.2)
bigWig files were generated using the bedGraphToBigWig tool. Score represents the normalized coverage of DNA fragments at a given genomic coordinate.
peaks were called by SECAR using IgG input as background and with the parameters of -norm -stringent -output
Assembly: MM10
Supplementary files format and content: bigwig
Supplementary files format and content: bed file for the peak
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| Submission date |
Jul 21, 2023 |
| Last update date |
Feb 29, 2024 |
| Contact name |
Martin M Matzuk |
| E-mail(s) |
mmatzuk@bcm.edu
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| Organization name |
Baylor College of Medicine
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| Department |
Pathology and Immunolodgy
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| Lab |
Matzuk
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| Street address |
One Baylor Plaza
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| City |
Houston |
| State/province |
TX |
| ZIP/Postal code |
77030 |
| Country |
USA |
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| Platform ID |
GPL19057 |
| Series (1) |
| GSE237975 |
Utilization of Tagged Transgenic Mouse Lines to Study the Molecular Roles of SMAD1/5 in Mediating Signaling Crosstalk During Early Pregnancy II |
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| Relations |
| BioSample |
SAMN36679652 |
| SRA |
SRX21126750 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7657202_S1HArep2_PR.bw |
83.6 Mb |
(ftp)(http) |
BW |
| GSM7657202_SMAD1-PR-rep2.bed.gz |
300.3 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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