|
| Status |
Public on Jul 10, 2023 |
| Title |
6hpa_control_H3K9me3_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
6hpa
|
| Organism |
Mus musculus |
| Characteristics |
treatment: wild type
chip antibody: H3K9me3 (Active Motif, 39161)
cell type: SCNT embryo 6 hpa
|
| Treatment protocol |
Three siRNAs against Suv39h2 were diluted and mixed in nuclease-free water to a working concentration of 20 μM per siRNA. Oocytes were injected with approximately 10 pL of siRNAs using a Piezo-driven micromanipulator. The injected oocytes underwent SCNT procedure after 20 min recovery. Embryos were collected at 6hpa for H3K9me3 CUT&RUN.
|
| Growth protocol |
MII oocytes were retrieved from the dissected oviducts of super-ovulated B6D2F1 female mice at 13 h post hCG injection for subsequent SCNT procedure. To get fertilized embryos, the super-ovulated C57BL/6 female mice were mated with C57BL/6 male mice. Then, the zygotes were collected from the oviducts of the female mice at 20 h after hCG injection and were cultured in G-1 PLUS medium at 37℃ under 5% CO2 until blastocysts.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
50 blastomeres were used per reaction. Each sample were bound with 5 μL concanavalin-coated magnetic beads and incubated with 1 μg antibody at 4℃ for 2 hours. Deluted pA-MNase was incubated with samples at 4℃ for 1 hour and chromatin cleavage was initiated with 100mM CaCl2,for 15 minutes at 0℃. RNA and proteins were then digested. Fragmented DNA was purified by phenol chloroform followed by ethanol purification.
Libraries were generated using the KAPA HyperPrep Kit for the Illumina platform (kk8504), following the manufacturer's instrunctions. Paired-end 150 bp sequencing was performed in NovaSeq (Illumina) platform in Berry Genomics and Novogen.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Low-quality and the adaptor sequences were trimmed from the reads using cutadapt (v1.16)with parameters: --trim-n -q 25,25 -m 75 -a AGATCGGAAGAGC -A AGATCGGAAGAGC.
Then, all CUT&RUN data reads were aligned to the mm10 reference genome used BWA with mem command.
Signal tracks were generated using the deepTools bamCoverage function with normalized to 1-million reads by sequencing depth of spike in.
Assembly: mm10
Supplementary files format and content: *.bw are bigWig files generated using deepTools.
Library strategy: CUT&RUN
|
| |
|
| Submission date |
May 15, 2023 |
| Last update date |
Jul 10, 2023 |
| Contact name |
Qianshu Zhu |
| E-mail(s) |
zhuqianshu@tongji.edu.cn
|
| Organization name |
Tongji University
|
| Department |
School of Life Science and Technology
|
| Street address |
Siping Road
|
| City |
Shanghai |
| State/province |
Shanghai |
| ZIP/Postal code |
270000 |
| Country |
China |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE194327 |
Unreprogrammed H3K9me3 prevents minor zygotic genome activation and lineage commitment in SCNT embryos |
| GSE195762 |
Unreprogrammed H3K9me3 prevents minor zygotic genome activation and lineage commitment in SCNT embryos |
|
| Relations |
| BioSample |
SAMN35099375 |
| SRA |
SRX20358258 |
