|
| Status |
Public on Jan 30, 2022 |
| Title |
NKO mice_snRNA-seq_NKO_male rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Hypothalamus (Tuberal and Mammillary zones)
|
| Organism |
Mus musculus |
| Characteristics |
strain background: C57BL/6
tissue: Brain
age: Postnatal day 14
genotype: NKO
tissue: Hypothalamus (Tuberal and Mammillary zones)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Lineage tracing experiment: Hypothalamus were dissected from Tbx3-CreER::Ai14 mice (E9 > P12). Then Tbx3-derived cells were isolated by FACS and sequenced by scRNA-seq. Tbx3 knockout experiment: Tuberal and mammillary zones of hypothalamus were dissected from NKO mice and its littermate control. Then tissue were dissociated into single cell for scRNA-seq, or were ground to power to extract the nuclei for snRNA-seq. CUT&Tag experiment: Hypothalamus were dissected from NKO mice and its littermate control and ground to power to extract the nuclei for CUT&Tag.
We took advantage of DNBSEQTM technology platforms (BGI Genomics, China) and DNBelab C4 Single-Cell Library Prep Set (MGI, #1000021082) for single-nucleus RNA library preparation. And we followed the manufacturer’s recommendations for the 10x Genomics 3’ Gene Expression v3 chemistry to prepare single-cell RNA library. For CUT&Tag library, DNA samples were processed with TruePrep DNA Library Prep kit (Vazyme; #TD501-01) according to manufacturer’s instructions.
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| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
DNBSEQ-T7 |
| |
|
| Description |
C_NKO mice snRNA-seq_neurons_50872.rds
|
| Data processing |
CellRanger Mkfatsq/Count Analysis
For scRNA-seq or snRNA-seq, the matrix files arising from different batches of experiments were subsequently integrated and analyzed using Seurat v3 software. For quality control, cells or nuclei with more than 10% mitochondrial gene counts were removed. In addition, cells with less than 600 genes or 500 UMIs were filtered out from the analysis, while nuclei were filtered on the basis of gene detection between 800 and 8,000.
CUT&Tag data were mapped to mouse (mm9) reference genome using Bowtie2 software. Only paired-end reads with both ends mapped correctly were kept for further analysis. All multimapped reads were removed using SAMtools for further analysis and duplicate reads were removed using Sambamba software. Peak calling was performed using peak calling algorithm MACS2.
Genome_build: mm10 for RNA-seq, mm9 for CUT&Tag
Supplementary_files_format_and_content: Lineage tracing experiment: A_TA lineage tracing_total cells_9224.rds; Tbx3 knockout experiment: B_NKO mice scRNA-seq_total cells_45406.rds, C_NKO mice snRNA-seq_neurons_50872.rds. RDS files are serialized R objects of the Bioconductor class ExpressionSet that contains the final data used to generate the results. It can be imported into R with `readRDS(“A_TA lineage tracing_total cells_9224.rds”)`. It contains the gene counts, sample metadata, and feature metadata for single cell / nucleus that passed quality control.
Supplementary_files_format_and_content: CUT&Tag experiment: D_02_89-Ab-WT_peak_summits.motif.bed, D_03_89-Ab-WT_peak_summits.motif.bed. BED is used to save genomic data and linked annotations in a text format. BED files can be read and modified using bedtools, Integrative Genomics Viewer (IGV) and UCSC genome browser.
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| |
|
| Submission date |
Jan 26, 2022 |
| Last update date |
Jan 30, 2022 |
| Contact name |
Qing-Feng Wu |
| E-mail(s) |
wu_qingfeng@genetics.ac.cn
|
| Organization name |
Institute of Genetics and Developmental Biology of the Chinese Academy of Sciences
|
| Street address |
No. 1 West Beichen Road, Chaoyang District
|
| City |
Beijing |
| ZIP/Postal code |
100101 |
| Country |
China |
| |
|
| Platform ID |
GPL28330 |
| Series (1) |
| GSE195472 |
Hierarchical deployment of Tbx3 dictating the identity of hypothalamic Tac2/Kiss1 neurons |
|
| Relations |
| BioSample |
SAMN25284857 |
| SRA |
SRX13934454 |
