 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Oct 26, 2021 |
| Title |
CL315-12hAUX-ESC-2 [bulk RNA-seq] |
| Sample type |
SRA |
| |
|
| Source name |
ESC
|
| Organism |
Mus musculus musculus x Mus musculus castaneus |
| Characteristics |
strain: C57Bl6/Cast
Sex: female
cell type: ESC
cell line: AID-tagged SPEN with dIDR-SPEN rescue
xist: tetO-Xist
differentiation: None
treatment: 12h Auxin
|
| Growth protocol |
ESCs were grown in 2i + LIF medium.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated using the Zymo Research RNA miniprep isolation kit (cat. #50-444-629).
RNA-seq libraries were prepared using the TrueSeq Stranded mRNA Library Prep Kit (Illumina 20020594).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
Bulk RNA-seq of SPEN-AID degron female ESCs with dIDR-SPEN rescue construct and tetOXist allele on Bl6 X chromosome, with 12h AUX treatment.
Mus musculus x Mus castaneous
mRNA-seq
processed data file: SPEN_WT_dIDR_rescue_featurecounts.txt
processed data file: spen_WT_KO_dIDR_rescue_bulk_ttest.csv
|
| Data processing |
[mRNA-seq]
Bulk RNA-seq data was aligned to the mm10 genome with STAR (v2.7.1a), filtered with samtools (v1.7) and allelic ratios were called using GATK (v4.1.4.1) ASEReadCounter.
Allelic reads were summed across genes. Genes with fewer than 10 allelic reads were filtered out from each sample. Allelic ratios between 0h and 24h dox were compared with a one-sided welch t-test (p<0.05), where each replicate is a sample.
[10X 3'GEX]
Cellranger (v5.0.1) was used to align and process the reads to the mm10 genome, Seurat (v3.9.9.9024) in R (v3.6.1) to filter out low quality cells, and vartrix (v1.1.14) to generate allelic ratios for each SNP in each cell.
Allelic reads were summarized across genes. Lowly expressed genes were filtered out, by requiring a minimum of 10 cells in each sample to have at least one allelic read expressed, and a minimum of 3 allelic reads per gene in each sample. Xist is induced by dox from the Bl6 allele, all cells were assumed to inactivate the Bl6 X chromosome. Allelic ratios between 0h and 24h dox for each mutant were compared using a one-sided welch t-test (p<0.05), where each cell is a sample.
[CLAP-seq]
CLAP-seq of full length, dIDR and dRRM SPEN was aligned to the mm10 genome with STAR (v2.7.1a), filtered with samtools (v1.7), duplicate reads were removed with GATK (v4.1.4.1) MarkDuplicates. RPKM was calculated for each sample in 100bp bins smoothed across 300bp with deeptools (v3.5.0), and enrichment was calculated by dividing SPEN pulldown RPKM over input RPKM in each bin.
Genome_build: mm10 (GRCm38)
Supplementary_files_format_and_content: SPEN_WT_dIDR_rescue_featurecounts.txt: featureCounts file.
Supplementary_files_format_and_content: spen_WT_KO_dIDR_rescue_bulk_ttest.csv: t-test results of 24h vs 0h allelic ratios in 3 replicates of WT, dIDR, and dSPOC.
Supplementary_files_format_and_content: features.tsv, barcodes.tsv, and matrix.mtx: Cellranger outputs.
Supplementary_files_format_and_content: spen_WT_dIDR_dSPOC_10x_ttest.csv: t-test results of 24h vs 0h allelic ratios in WT, dIDR, and dSPOC single cells.
Supplementary_files_format_and_content: DeltaB_WT_D2_ttest.csv: t-test results of deltaB vs WT allelic ratios in 129-Xist expressing single cells at day 2 of differentiation.
Supplementary_files_format_and_content: DeltaB_WT_D4_ttest.csv: t-test results of deltaB vs WT allelic ratios in 129-Xist expressing single cells at day 4 of differentiation.
Supplementary_files_format_and_content: dIDR_A.rpkm.Xist.bedgraph: RPKM of dIDR-SPEN binding to Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: dIDR_B.rpkm.Xist.bedgraph: RPKM of dIDR-SPEN binding to Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: dRRM_B.rpkm.Xist.bedgraph: RPKM of dRRM-SPEN binding to Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: FL_A.rpkm.Xist.bedgraph: RPKM of FL-SPEN binding to Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: FL_B.rpkm.Xist.bedgraph: RPKM of FL-SPEN binding to Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: INPUT_dIDR_A.rpkm.Xist.bedgraph: RPKM of dIDR-SPEN input over Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: INPUT_dIDR_B.rpkm.Xist.bedgraph: RPKM of dIDR-SPEN input over Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: INPUT_dRRM_B.rpkm.Xist.bedgraph: RPKM of dRRM-SPEN input over Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: INPUT_FL_B.rpkm.Xist.bedgraph: RPKM of FL-SPEN input over Xist RNA in 100bp bins smoothed across 300bp.
Supplementary_files_format_and_content: INPUT_FL_A.rpkm.Xist.bedgraph: RPKM of FL-SPEN input over Xist RNA in 100bp bins smoothed across 300bp.
|
| |
|
| Submission date |
Jul 30, 2021 |
| Last update date |
Oct 26, 2021 |
| Contact name |
Kathrin Plath |
| Organization name |
University of California, Los Angeles
|
| Street address |
615 Charles E Young Dr S
|
| City |
Los Angeles |
| State/province |
CA |
| ZIP/Postal code |
90095 |
| Country |
USA |
| |
|
| Platform ID |
GPL30463 |
| Series (2) |
| GSE181235 |
Xist-seeded nucleation sites form local concentration gradients of silencing proteins to inactivate the X-chromosome [II] |
| GSE181236 |
Xist-seeded nucleation sites form local concentration gradients of silencing proteins to inactivate the X-chromosome |
|
| Relations |
| BioSample |
SAMN20507248 |
| SRA |
SRX11613873 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
|
|
|
|
 |
