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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jan 24, 2021 |
| Title |
Ythdc1_cKO_ESC#1_+4OHT.rep1 |
| Sample type |
SRA |
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| Source name |
embryonic stem cells
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| Organism |
Mus musculus |
| Characteristics |
cell type: embryonic stem cells
genotype: Ythdc1_cKO
esc id: 1
treatment: 4OHT
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| Growth protocol |
Male mouse embryonic stem cells (mESCs) were derived from 3.5 d.p.c inner cell mass (ICM) from female 129 mice (Stock No.: 217, Beijing Vital River Laboratory Animal Technology Co., Ltd.). Animal care and experimental protocols were approved by the animal ethics committee of the Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences. ESCs with genetic engineering, including Ythdc1-conditional knockout (cKO), MERVL::tdTomato reporter, and Halo- Ythdc1 knockin (KI) , and TurboID overexpression (OE) were constructed and cultured in feeder- free condition, using serum/LIF medium containing high-glucose Dulbecco’s modified Eagle’s medium (DMEM, HyClone, #SH30022.01), 15% fetal bovine serum (FBS, Front Biomedical, # OPT500), 1× GlutaMAX, 1× nonessential amino acids (NEAA, Gibco, #11140076), 1× Penicillin- Streptomycin (Hyclone, #SV30010), 1× Sodium Pyruvate (Gibco, #11360070), 1 mM 2- Mercaptoethanol (Gibco, # 21985023), and 1000 U/ml LIF. HEK293T (ATCC, # CRL-1126) and NIH3T3 (ATCC, #CRL-1658) cells were maintained in high-glucose DMEM supplemented with 10% FBS (Natocor, #SFBE), 1× GlutaMAX (Gibco, #35050079), and 1× NEAA. All the cell lines were cultivated at 37°C in a humidified atmosphere containing 5% CO2.
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| Extracted molecule |
total RNA |
| Extraction protocol |
total RNA
Total RNAs were extracted as described above. The VAHTS Total RNA-seq (H/M/R) Library Prep Kit for Illumina (VAHTS, #NR603) was used for RNA library preparation. In brief, 1 μg total RNAs were hybridized with the rRNA probe (H/M/R) and digested by RNase H to remove ribosomal RNAs. After DNase I digestion, the ribosomal-depleted RNAs were fragmented at 94°C for 8min. Then the first-strand and second-strand cDNAs were synthesized successively using the provided reagents. The cDNA was purified by VAHTS DNA Clean Beads (VAHTS, #NR411), followed by end repaired, dA-tailing, adapter ligation, and second strand cDNA digestion. After two-round of purification, the cDNAs was PCR-amplified and purified by VAHTS DNA Clean Beads. High- throughput sequencing was performed using Illumina HiSeq Xten (Illumina) platform with PE150 (pair-end sequencing, 150 bp reads).
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
HiSeq X Ten |
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| Description |
Processed data file: GSE146466_RNA-seq-expr.tsv.gz
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| Data processing |
bcl2fastq conversion software v2.17 used for basecalling.
Reads were aligned to the gencode (mm10 vM21) transcriptome using STAR (version 2.7)
The gene expression level was calculated by using STAR
The RNA-seq data processing was performed as described45. To analyze the regular gene expression, the reads were aligned to the reference transcriptome by RSEM46 and bowtie2 (—bowtie2— bowtie2-sensitivity-level very_sensitive —no-bam-output —estimate -rspd) using the index built by RSEM with the mouse genome, mm10, and Ensembl gene annotation track v74
Genome_build: mm10
Supplementary_files_format_and_content: expression matrx
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| Submission date |
Mar 05, 2020 |
| Last update date |
Jan 24, 2021 |
| Contact name |
Jiangping He |
| E-mail(s) |
he_jiangping@grmh-gdl.cn
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| Organization name |
Guangzhou Institutes of Biomedicine and Health (GIBH), CAS
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| Street address |
Kai yuan avenue 190
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| City |
GuangZhou |
| ZIP/Postal code |
510530 |
| Country |
China |
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| Platform ID |
GPL21273 |
| Series (2) |
| GSE146466 |
High throughput sequencing for YTHDC1 binding RNAs [RNA-seq] |
| GSE146467 |
The RNA m6A reader YTHDC1 silences retrotransposons and guards ES cell identity |
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| Relations |
| BioSample |
SAMN14307354 |
| SRA |
SRX7857095 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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