|
| Status |
Public on Oct 19, 2023 |
| Title |
CTRi_L2CRNA_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Late 2-cell embryo
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Late 2-cell embryo
treatment: dCas9-KRAB mRNA + control sgRNA
|
| Treatment protocol |
About 6 pl dCas9-KRAB mRNA and sgRNA mixes were injected into cytoplasm of zygotes between 2-4 hpi using a Piezo impact-driven micromanipulator (Eppendorf). For all micro-injection experiments, sgRNAs were injected at 60ng/µl: 10ng/µl for each MT2_Mm sgRNA or 60ng/µl for the non-targeting control sgRNA. For embryos used for total RNA-seq analyses, dCas9-KRAB mRNA was injected at 100ng/µl.
|
| Growth protocol |
Embryos were cultured in KSOM (Millipore) at 37 °C under 5% CO2 with air
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Embryos were suspended in 7 μl RRI water (199 μl ultrapure H2O with 1 μl RNase inhibitor). Then, 6 μl shearing master mix (1 μl 10X lysis mix, 1 μl Smart scN6, 4 μl scRT Buffer) was added and incubated at 85°C for 6 min and immediately put on ice for 2 min.
The total RNA‐seq libraries were generated using the SMART‐Seq Stranded Kit (Takara) following the manufacturer's protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Raw RNA-seq read pairs were trimmed by Trimgalore (v0.6.6)(https://github.com/FelixKrueger/TrimGalore) with the option “--paired --clip_R2 5”, and aligned to mm10 by STAR (v2.7.9) with the options “--outFilterMultimapNmax 100 --winAnchorMultimapNmax 100 --outSAMattributes NH HI NM MD XS AS --alignIntronMax 1000000 --alignMatesGapMax 1000000 --limitBAMsortRAM 30000000000 --outFilterType BySJout”.
Uniquely aligned read pairs were identified by sambamba (v0.6.8) with the option “-F “mapping_quality >=255”
Bigwig files were generated using deeptools (v2.0.0) bamCoverage function with the options --binSize 25 --normalizeUsing RPKM --scalFactor 1”
Assembly: mm10
Supplementary files format and content: Bigwig
|
| |
|
| Submission date |
Aug 31, 2023 |
| Last update date |
Oct 19, 2023 |
| Contact name |
Zhiyuan Chen |
| E-mail(s) |
[email protected]
|
| Organization name |
Cincinnati Children's Hospital Medical Center
|
| Street address |
3333 Burnet Avenue
|
| City |
Cincinnati |
| State/province |
Ohio |
| ZIP/Postal code |
45229 |
| Country |
USA |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE242121 |
Systematic Perturbation of Thousands of Retroviral LTRs in Mouse Embryos [RNA-Seq] |
| GSE242123 |
Systematic Perturbation of Thousands of Retroviral LTRs in Mouse Embryos |
|
| Relations |
| BioSample |
SAMN37220578 |
| SRA |
SRX21586191 |
