|
| Status |
Public on May 11, 2022 |
| Title |
scRNA-seq_E12-5_XX_ECKO_Embryo_1_Rep_11 |
| Sample type |
SRA |
| |
|
| Source name |
Primordial germ cells
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Primordial germ cells
strain: C57BL/6;129S1/SvlmJ
developmental stage: E12.5
genotype: Eed -/-
gender: Female
|
| Growth protocol |
Control and ECKO male and female embryos were obtained from crosses between OG; Eedfl/fl (Lengner et al., 2007; Yu et al., 2009) homozygous females and BC; Eedf/l+ heterozygous males (Ohinata et al., 2005) at E10.5, E11.5 and E13.5. Embryos were staged by the detection of a vaginal plug on the morning after time-mating pairs were established E0.5.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Female E12.5 gonads were extracted and digested in 0.05% trypsin. GFP positive single mPGCs were sorted into a 96 well skirtless plate, each well containing 3 µL of 0.2% Triton X-100 and 2U/µL RNaseOUT (Thermo Fisher Scientific).
Single Cell RNA sequencing libraries were prepared according to the Smart Seq 2 single cell sequencing protocol (Picelli et al., 2014). The RNA was converted to cDNA in each well using Superscript II (Invitrogen). The cDNA was amplified in each well using a KAPA HiFI HotStart polymerase (KAPA Biosystems) in a PCR run for 18 cycles. cDNA was tagged via the Nextera Tagmentation kit (Illumina) using libraries A and C (Illumina). Indexed cDNA was pooled and run on a 2% agarose gel and the smear between 200-500 bp was extracted. Gel extract was cleaned with the Qiagen Gel Extraction Kit (Qiagen) and sent for sequencing.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
MattLowe5704_female_counts.txt
|
| Data processing |
The RNA-seq reads were aligned using HISAT2
Genes that are expressed (read counts > 0) in at least one replicate in both control and ECKO are included for analysis.
Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated using edgeR
WGBS reads were aligned with BS Seeker 2. CG sites with read depth >= 4 are preserved in the Cgmap.gz file.
Genome_build: mm9
Supplementary_files_format_and_content: Excel files includes RPKM values for each sample.
|
| |
|
| Submission date |
Aug 05, 2021 |
| Last update date |
May 11, 2022 |
| Contact name |
Pao-Yang Chen |
| Organization name |
Academia Sinica
|
| Department |
Institute of Plant and Microbial Biology
|
| Lab |
Pao-Yang Chen
|
| Street address |
128 Sec. 2, Academia Rd, Nankang,
|
| City |
Taipei |
| ZIP/Postal code |
115 |
| Country |
Taiwan |
| |
|
| Platform ID |
GPL24247 |
| Series (1) |
| GSE139413 |
EED is required for Mouse Primordial Germ Cell Differentiation in the Embryonic Gonad |
|
| Relations |
| BioSample |
SAMN20603778 |
| SRA |
SRX11664804 |
