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Sample GSM5849748 Query DataSets for GSM5849748
Status Public on Jul 10, 2023
Title NT-ICM_RNA_rep5
Sample type SRA
 
Source name NT-ICM
Organism Mus musculus
Characteristics development stage: ICM
treatment: untreated
cell type: NT
Treatment protocol Isolated zygotes were injected with either morpholino oligonucleotides targeted at Max, Mcrs1 or 25-base random control morpholino. All morpholinos were designed and synthesized in Gene Tools (https://www.gene-tools.com/). Injected embryos were cultured in G-1 PLUS medium at 37℃ under 5% CO2. Knocking-down (KD)-efficiency was examined at morula or blastocyst stage. ICM and TE were collected for Smart-seq2.
Growth protocol BDF1 mice were used as oocyte donors, and the cumulus cells were used as nuclear donors. MII oocytes were enucleated in Chatot-Ziomek-Bavister (CZB) medium containing 5 μg/mL cytochalasin B (CB) by Piezo-driven pipette (PrimeT 130 ech) of an Olympus inverted microscope. The donor cells were transferred into enucleated oocytes by direct injection. The reconstructed oocytes were then cultured in CZB medium for 1h before activation treatment. The cloned constructs were activated by 6 h incubation in 1 mM SrCl2 in Ca2+-free CZB and 5 μg/mL cytochalasin B. Reconstructed embryos were thoroughly washed and cultured in G-1 PLUS medium at 37℃ under 5% CO2.
Extracted molecule polyA RNA
Extraction protocol 10 blastomeres were used per reaction, and three replicates were performed for each stage. All isolated blastomeres were washed three times in 0.5% BSA in PBS solution to avoid contamination. RNA-seq libraries were generated using the Covaris DNA shearing protocol for Smart-seq sequence library generation. Briefly, RNAs with a polyadenylated tail were captured, reverse transcribed and pre-amplified. DNA was sheared and purified for library construction.
Libraried 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 RNA-Seq
Library source transcriptomic
Library selection cDNA
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.
All RNA-seq data were mapped to the mm10 reference genome after cutadapt using hisat2 with parameters ‘--data-cufflinks --no-discordant --no-mixed --no-unal’.
The TPM (Transcripts Per Million) of genes were obtained by StringTie.
Signal tracks for each sample were generated using the deepTools bamCoverage function with parameters "--normalizeUsing RPKM --smoothLength 150 --extendReads 150".
Genome_build: mm10
Supplementary_files_format_and_content: *.bw are bigWig files generated using deepTools. *.exp are gene expression files generated by StringTie.
 
Submission date Jan 31, 2022
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)
GSE195760 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 SAMN25512645
SRA SRX13993291

Supplementary file Size Download File type/resource
GSM5849748_NT-ICM_rep5.bw 13.6 Mb (ftp)(http) BW
GSM5849748_NT-ICM_rep5.exp.gz 1.1 Mb (ftp)(http) EXP
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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