|
| Status |
Public on Dec 17, 2021 |
| Title |
Hk2 fl/fl repopulated microglia |
| Sample type |
SRA |
| |
|
| Source name |
Brain
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
genotype: Hk2 fl/fl
cell type: isolated CD11b high CD45 low cells
|
| Treatment protocol |
For induction of the Cre-dependent recombination in postnatal mice, 100 μg tamoxifen (TAM, Sigma) dissolved in corn oil was injected intraperitoneally for 2 consecutive days at postnatal day 1-2 or as indicated.
|
| Growth protocol |
Cx3cr1CreER mice were purchased from the Jackson Laboratory. hk2 floxed (hk2 fl/fl) mice and Hk2CreER-tdTomato mice were generated by Biocytogen Company (China). hk2 fl/fl mice were crossed with the Cx3cr1CreER/+mice to generate the Cx3cr1CreER::hk2 fl/fl or hk2 fl/fl animals. All animals were housed under a 12-h light/dark cycle with food and water available.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Mice were anesthetized with sodium pentobarbital and perfused with ice-cold 0.9% NaCl solution. Brains were rapidly dissected and transferred to a pre-chilled dish with cold PBS on ice. The brains were minced and dissociated in DMEM medium (Fischer Scientific) containing 0.5 mg/ml collagenase IV (Sigma) and 10 U/ml DNase I (Roche). Myelin and tissue debris were removed by centrifuging through a 37-70% (v/v) Percoll gradient (GE Healthcare BioSciences) density. Mononuclear cells were collected and incubated with different antibodies as indicated, and microglia were purified as a CD11b+CD45low populations on a BD FACSAria II using the 80 μm micron nozzle.
Single-cells were captured and libraries were prepared following the manufacturer’s instructions of 10x Chromium Next GEM Single Cell 3' GEM and Library & Gel Bead Kit v3.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Libraries were sequenced on an Illumina NovaSeq 6000 sequencing system (paired-end multiplexing run, 150bp) by LC-Bio Technology Co. Ltd., (HangZhou, China) at a minimum depth of 20,000 reads per cell.
Raw sequencing data demultiplexing and converting were carried out by Illumina bcl2fastq software.
Sample demultiplexing, barcode processing and single-cell 3’ gene counting were included in the preprocessing steps of Cell Ranger pipeline (version 3.1.0).
Data were aligned to Ensembl genome GRCm38 reference genome.
Genome_build: GRCm38
Supplementary_files_format_and_content: Expression matrices output by Cellranger are provided as mtx files. The gene and cell barcode name files that accompany the mtx files are provided as tsv files (.tsv) for each sample.
|
| |
|
| Submission date |
Nov 11, 2021 |
| Last update date |
Dec 17, 2021 |
| Contact name |
weiying wu |
| E-mail(s) |
wuweiying@zju.edu.cn
|
| Phone |
19883131534
|
| Organization name |
Zhejiang University
|
| Street address |
866 Yuhangtang Rd
|
| City |
Hangzhou |
| State/province |
Zhejiang |
| ZIP/Postal code |
310027 |
| Country |
China |
| |
|
| Platform ID |
GPL24247 |
| Series (1) |
| GSE188642 |
Single-cell RNA sequencing of repopulated microglia at day 3 |
|
| Relations |
| BioSample |
SAMN22488070 |
| SRA |
SRX12753106 |
