 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Apr 14, 2024 |
| Title |
SPATAC-seq with mouse organogenesis well-E1G |
| Sample type |
SRA |
| |
|
| Source name |
mouse embryo
|
| Organism |
Mus musculus |
| Characteristics |
tissue: mouse embryo
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Female mice were mated naturally, and the first day that the vaginal plug was observed was considered as E0.5. Dissections were performed as previously described (Cao J, et al. Nature. 2019) and all embryos were immediately snap-frozen in liquid nitrogen. Frozen embryos were stored at −80 °C. Embryo nuclei were extracted via previously described methods with some modifications (Corces MR, et al. Nat Methods. 2017; Cao J, et al. Nature. 2019). Briefly, each embryo was transferred into a pre-chilled 2 ml Dounce (D8938, Sigma) with 2 ml Homogenization Buffer (10 mM Tris-HCl pH 7.8, 5 mM CaCl2 (21115, Sigma), 3 mM Mg (Ac)2 (63052, Sigma), 0.1% NP-40 (11332473001, Sigma), 0.1mM EDTA (AM9260G, Thermo), 320 mM Sucrose (S8271-500G, Solarbio), 0.015mM PMSF (P7626, Sigma) and 0.15mM β-mercaptoethanol (M3148, Sigma)), and homogenized by pipetting 10 times with A pestle and B pestle, respectively. Homogenized embryos were filtered with a 40 μm filter. The filtered nuclei were then transferred to a new 15-ml tube containing 10ml nuclei wash buffer (NWB) (10 mM Tris-HCl pH 7.4, 10 mM NaCl (AM9759, Thermo), 3 mM MgCl2 (AM9530G, Thermo), 1% BSA (130-091-376, Miltenyi Biotec) and 0.1% Tween-20 (11332465001, Sigma)) and pelleted by spinning 5 min at 4°C at 350 g and washed once with 10ml NWB. The nuclei were resuspended in 10ml NWB and fixed by adding 140 μl of 37% formaldehyde for exactly 10 min on ice. The fixation was stopped by adding 500 μl of 2.5M glycine (G8898, Sigma). The sample was incubated at room temperature for 5 minutes and then centrifuged at 500 g for 5 minutes to remove supernatant. The nuclei pellet was washed twice with 10 ml NWB and re-suspended in 2 ml NWB. Nuclei were then counted on a hemocytometer and diluted to 5,000 nuclei per 1 μl using NWB. Finally extracted nuclei from all samples were loaded into SPATAC-seq, a custom high-throughput single-cell ATAC-seq method, to capture the accessible regions during mouse organogenesis, which profiling more than 360,000 cells in one experiment.
Bulk ATAC-seq and SPATAC, a customized single-cell ATAC-seq library construction method, see description of our paper for details.
|
| |
|
| Library strategy |
ATAC-seq |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Raw Read1 Fastq files within genome DNA (position from 66-121 nucleotide) were extracted and renamed as new Read1 file. Similarly, raw Read2 Fastq files within genome DNA (position from 51-101 nucleotide) were extracted and renamed as new Read2 file of Cell Ranger ATAC format. Round 1, Round2 and Round3 barcodes are extracted from raw Read1 and Read2 Fastq files, are merged after correcting 1bp mismatch. Then merged barcodes were attached in the read name of new Read1 and Read2 fastq files by software sinto (https://timoast.github.io/sinto/index.html), which were aligned to the reference mouse (mm10) genome (https://cf.10xgenomics.com/supp/cell-atac/refdata-cellranger-atac-mm10-1.2.0.tar.gz) using BWA. The bam file of each sublibrary was converted to scATAC-seq fragment file by sinto according to standard procedure. Finnaly, the barcodes in fragment file of each sublibrary were tagged with Round4 barcodes (PCR wells).
Assembly: mm10
Supplementary files format and content: bed files, the fragment file of SPATAC-seq, the information of each unique fragment of SPATAC within each sublibrary, including the chromsome number, start site, end site and frequency.
Supplementary files format and content: CSV file, cell metadata information for the atlas of zebrafish development, including cell types, coordinate and quality metric of each cell
|
| |
|
| Submission date |
Oct 23, 2022 |
| Last update date |
Apr 14, 2024 |
| Contact name |
Keyong Sun |
| E-mail(s) |
keyongsuntsinghua@gmail.com
|
| Organization name |
tsinghua university
|
| Department |
School of Medicine
|
| Street address |
Hai Dian street
|
| City |
beijing |
| ZIP/Postal code |
100084 |
| Country |
China |
| |
|
| Platform ID |
GPL24247 |
| Series (1) |
| GSE216371 |
A single-cell atlas of chromatin accessibility in mouse organogenesis |
|
| Relations |
| BioSample |
SAMN31420821 |
| SRA |
SRX17998883 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6671139_E1G.bed.gz |
1001.5 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
