|
| Status |
Public on Jul 10, 2024 |
| Title |
inflamed_CCR2_rep3 |
| Sample type |
SRA |
| |
|
| Source name |
Bone
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Bone
Sex: female
cell type: monocytes
treatment: inflamed
|
| Growth protocol |
Wild type and REP(13) mice were housed in a specific pathogen free animal facility at the University of Glasgow. All animal experimentation was carried out under the auspices of a UK Home Office licence and all procedures were approved by the local University of Glasgow ethics committee. All mice used were female between the ages of 8-12 weeks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cell suspensions were stained for 20 minutes at 4°C with 100 μL of fixable viability stain (eBioscience) and washed in FACS buffer (PBS containing 2mM EDTA and 2% FBS). Next, cells were stained for 20min at 4°C with 50 μL of subset-specific antibody cocktails (Supplementary Table 1) and washed in FACS buffer. For flow cytometry analysis, stained cells were fixed for 20min at 4°C in 100 μL of Fixation Buffer (BioLegend) and analysed on a BD LSRFortessa flow cytometer (BD Biosciences). For monocyte isolation and transcriptomic analysis, stained cells (Supplementary Table 2) were analysed on a FACS Aria sorter (BD Biosciences) without previous fixation. Monocytes expressing either CCR1/2 or CCR2 only were sorted in RLT buffer (Qiagen) containing 10 μL/mL of b-mercaptoethanol and stored at -80oC for RNA extraction.
RNA from sorted monocytes was isolated using the RNeasy® Micro Kit (Qiagen) as per manufacturer’s instructions. Next, mRNA libraries were prepared using the NEBNext® Single Cell/Low Input RNA Library Prep Kit for Illumina® (New England Biolabs).
Paired-end sequencing was performed in a NextSeq2000 sequencing platform (Illumina) aiming for 40 million reads sequencing depth.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 2000 |
| |
|
| Data processing |
Data was aligned to the mouse genome (GRCm38.91) using STAR (2.7.10a). The index was built for an overhang of 74 bp (read length -1).
Read counts were produced via STAR, under default settings.
Counts were combined into a single read count table using R.
Assembly: GRCm38.91
Supplementary files format and content: Raw read counts produced by alignment of each library to GRCm38 v91.
|
| |
|
| Submission date |
Dec 20, 2023 |
| Last update date |
Jul 10, 2024 |
| Contact name |
John Joseph Cole |
| Organization name |
University of Glasgow
|
| Street address |
University Avenue
|
| City |
Glasgow |
| ZIP/Postal code |
G128QQ |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL30172 |
| Series (2) |
| GSE251646 |
CCR1 and CCR2 co-expression on monocytes is nonredundant and delineates a distinct monocyte subpopulation. [RNA-Seq] |
| GSE251648 |
CCR1 and CCR2 co-expression on monocytes is nonredundant and delineates a distinct monocyte subpopulation. |
|
| Relations |
| BioSample |
SAMN38976091 |
| SRA |
SRX22974656 |
