|
| Status |
Public on Jun 04, 2024 |
| Title |
Sample_18_Neutrophil_APOE3_4_CN |
| Sample type |
SRA |
| |
|
| Source name |
Neutrophil
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Neutrophil
genotype: APOE3_4
disease state: CN
Sex: F
age: 56.5
qrds: 0
mmse: 30
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Blood was drowned in CPT tubes (BD Bioscience) and centrifuged at 1700g for 30min, to pellet polymorphonuclear leukocytes (neutrophils) below the gel barrier. Following the removal of plasma and peripheral blood mononuclear cells (PBMCs) from the fraction above the gel barrier, a syringe attached to an 18-gauge needle was used to extract the erythrocyte/neutrophil fraction below the gel as previously described. Cells collected in a 50ml tube were washed in PBS and centrifuged at 600g for 5min at 4°C. Cell pellet was resuspended twice in 10ml of ACK (ammonium chloride potassium) to lyse erythrocytes at RT and washed with PBS. Cells were stained with FITC anti-CD66b (1:100, Biolegend, 305103) in PBS supplemented with 2% fetal bovine serum (FBS) for 20min at 4°C. After staining, cells were washed and sorted using BD FACSAriaTM II (BD Bioscience).
Smart-Seq2 libraries were prepared by the Broad Technology Labs and sequenced by the Broad Genomics Platform. cDNA libraries were generated from sorted cells using the Smart-seq2 protocol. RNA sequencing was performed using Illumina NextSeq500 using a High Output v2 kit to generate 2× 25bp reads.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Count files (fastq) were preprocessed for quality control and adapter trimming using fastp. High quality reads were aligned using Salmon (v1.7).
Technical outliers were removed for further analysis. Downstream differential gene expression analysis was carried out using DESeq2 (v.1.34.00). Biological outliers were determined using PCA and heatmap visualization and removed for final analysis.
Comparisons were run using LRT, and the cutoff for significant genes was P < 0.05.
Assembly: GRCh38
Supplementary files format and content: zip file of salmon outputs including plain text quantification file (quant.sf), transcripts used for quantification, parameter settings, version of Salmon, and estimated bias parameters.
|
| |
|
| Submission date |
Sep 21, 2023 |
| Last update date |
Jun 04, 2024 |
| Contact name |
Neta Rosenzweig |
| E-mail(s) |
NROSENZWEIG@BWH.HARVARD.EDU
|
| Organization name |
Brigham and Women's Hospital, Harvard Medical School
|
| Street address |
60 Fenwood Road, 10002K, Brigham and Women’s Hospital
|
| City |
Boston |
| State/province |
Massachusetts |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (2) |
| GSE243743 |
SEX-DEPENDENT APOE4 NEUTROPHIL-MICROGLIA INTERACTIONS DRIVE COGNITIVE IMPAIRMENT IN ALZHEIMER'S DISEASE [Human_F_M_HC_MCI_AD_E33_34] |
| GSE243750 |
SEX-DEPENDENT APOE4 NEUTROPHIL-MICROGLIA INTERACTIONS DRIVE COGNITIVE IMPAIRMENT IN ALZHEIMER'S DISEASE |
|
| Relations |
| BioSample |
SAMN37502955 |
| SRA |
SRX21853422 |
