|
| Status |
Public on Sep 23, 2021 |
| Title |
Lung PDGFR-β+ cells scrRNA rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Lung PDGFR-β+ cells
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
tissue: Lung
cell type: PDGFR-Beta+ cells
age: 12-weeks old
genotype: WT
treatment: Klf4 scr RNA
|
| Treatment protocol |
Tracheal Smooth Muscle Cells were obtained from CellBiologics C57-6082 and maintained following cell culture indications
|
| Growth protocol |
Lungs were finely minced and incubated in 2 mg/ml collagenase in PBS at 37oC for 20 min. The digestion mixture was passed through a 14-gauge pipetting needle (Cadence Science), incubated at 37oC for an additional 20 min and then filtered through a cell strainer (Falcon). This single cell suspension filtrate was centrifuged at 1200 rpm for 5 min, and the pellet was resuspended in cold PBS with 1% fetal bovine serum (FBS; Invitrogen). The resuspended cells were incubated with either PE-conjugated anti-PDGFR-b antibody (1:200, Miltenyi Biotec) for 20 min in the case of Pdgfrb-CreERT2, Klf4(flox/flox) mice or DAPI (1:1000) to label dead cells for 10 min in the case of Pdgfrb-CreERT2, ROSA26R(Zs/+) mice. The sample was then washed in cold PBS with 1% FBS and centrifuged at 1200 rpm for 5 min. The resulting pellet was resuspended in PBS containing 1% FBS and 0.02% EDTA. DAPI (1:2000) was added to the cells that had undergone incubation with anti-PDGFR- antibody. Finally, PDGFR-+DAPI- or Zs+DAPI- cells were sorted on a BD FACSAria II cell sorter. Cells stained with DAPI alone were used as a control for anti-PDGFR- antibody specificity. Lungs from C57BL/6 mice were processed similarly to those of Pdgfrb-CreERT2, ROSA26R(Zs/+) mice, and isolated cells were used as a control for auto-fluorescence in the Zs channel.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
tpm-values-all-samples_PDGFR.txt
C2
|
| Data processing |
HISAT2 was used for alignment to the reference genome mm10
GENCODE annotations for mouse
ballgown/stringTie was used to generate the gene counts and transcript abundance estimates from the alignments.
DESeq2 was used for differential expression analysis using R
Genome_build: mm10
Supplementary_files_format_and_content: Matrix table with raw gene counts for every gene and every sample
|
| |
|
| Submission date |
Sep 23, 2021 |
| Last update date |
Sep 25, 2021 |
| Contact name |
Rolando Garcia-Milian |
| E-mail(s) |
rolando.milian@yale.edu
|
| Organization name |
Yale University
|
| Department |
CWML
|
| Lab |
Bioinformatics Support Hub
|
| Street address |
333 Cedar St
|
| City |
New Haven |
| State/province |
CT |
| ZIP/Postal code |
06510 |
| Country |
USA |
| |
|
| Platform ID |
GPL13112 |
| Series (2) |
| GSE184670 |
Mesenchymal cell type-specific roles of KLF4 in lung fibrosis [bulk RNA-seq] |
| GSE184672 |
Mesenchymal cell type-specific roles of KLF4 in lung fibrosis |
|
| Relations |
| BioSample |
SAMN21572670 |
| SRA |
SRX12310958 |
