|
| Status |
Public on Jun 07, 2023 |
| Title |
E2C_KO_Ribo1 |
| Sample type |
SRA |
| |
|
| Source name |
Early 2-cell embryos
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Early 2-cell embryos
strain: B6D2F1
developmental stage: Early 2-cell
female mice donor genotype: Eif4e1b KO; Rpl22 HA
|
| Extracted molecule |
other |
| Extraction protocol |
~200 oocytes or zygote, or ~100 early 2-cell embryos were used as one group for low input ribo-seq. Collected samples were washed with PBS and transferred into 1.5 ml nuclease free centrifuge tubes with a minimal volume of PBS. The tubes were frozen immediately in dry ice and stored at -80 until use.
The EZ-Manga RIP kit (Millipore 17-701) and anti-HA beads (Pierce 88836) were used for the low input ribo-seq experiment. Samples were first lysed following the RIP kit manual and then 100U RNase I (Ambion AM2294) was added to each tube for RNA digestion. The digestion was done at room temperature for 45 min. After digestion, anti-HA magnetic beads were added to each tube to finish IP at 4 degrees for 4h following the manual. 7ul ultrapure water supplied with 0.25 ul RNase inhibitor from the SMARTer smRNA-Seq Kit (Takara 635029) was added to each tube and the tubes were heated for 15min at 70 degrees to denature the proteins and RNAs inside. The tubes were transferred to ice immediately after heating and the remaining part of the manual was followed to finish library construction. 22 PCR cycles were used for indexing of libraries and the finial PCR products were purified with AMPure XP beads (Bechman A63881) for 2 time at 1:1.5 ratio.
|
| |
|
| Library strategy |
OTHER |
| Library source |
other |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
Ribosome protected RNA fragments, S11
|
| Data processing |
Base calling was finished with RTA2.
Sequenced reads were trimmed using cutadapt by indicating “AAAAAAAAAA” as the adapter sequence file, then mapped to the reference genome using STAR ver. 2.7.6a. The reference contained annotation of GRCm38 and rDNA.
Bam files containing only uniquely mapped reads were extracted and StringTie ver. 2.1.4 was used to generate counts of genes in the reference. deepTools ver. 3.5.1 was used to normalize the bam files by RPKM and generate the bigWig files.
Assembly: GRCm38
Supplementary files format and content: Tab-delimited text files include FPKM, TPM values for each sample.
Supplementary files format and content: bigWig file of uniquely mapped reads for each sample.
Supplementary files format and content: Matrix table with raw gene counts for every gene and every sample
Library strategy: Ribo-seq
|
| |
|
| Submission date |
Apr 18, 2023 |
| Last update date |
Jun 07, 2023 |
| Contact name |
Guanghui Yang |
| E-mail(s) |
guanghui.yang@nih.gov
|
| Organization name |
National Institutes of Health
|
| Department |
National Institute of Diabetes and Digestive and Kidney Diseases
|
| Lab |
Laboratory of Cellular and Developmental Biology
|
| Street address |
50 South Drive
|
| City |
Bethesda |
| State/province |
Maryland |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL24247 |
| Series (2) |
| GSE180218 |
Germ cell-specific eIF4E1b regulates maternal mRNA translation to ensure zygotic genome activation |
| GSE230019 |
Germ-cell specific eIF4E1b regulates maternal mRNA translation to ensure zygotic genome activation [Ribo-seq] |
|
| Relations |
| BioSample |
SAMN34239068 |
| SRA |
SRX20004188 |
