|
| Status |
Public on May 14, 2023 |
| Title |
MLL7 PDX AZD1775 (Wee1i) scRNAseq |
| Sample type |
SRA |
| |
|
| Source name |
frozen bone marrow cells
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: frozen bone marrow cells
disease state: KMT2A-r acute lymphoblastic leukemia
genotype: KMT2A::AFF1;MEF2D::PTMA; KMT2A::PTPRC and clonal mutations in FLGE3292G; FLT3I836del; METL575I; CHEK1I465T; CSF1RS422C
time: 28 h
construct/treatment: AZD1775
|
| Treatment protocol |
Mice were treated for 24 h with AZD1775 (Wee1i) (HY-10993 MedChemExpress) or DMSO (Sigma-Aldrich) when 1% of hCD45pos(BD Bioscience) hCD19pos (Biolengend) mCD45neg (Biolengend) cells were detected in peripheral blood. 4 hours after the second dose, mice were sacrificied by cervical dislocation and tibia, femur, and hip from both legs were collected and homogenized into single-cell suspensions by manual trituration and viably frozen in 10% DMSO (Sigma-Aldrich)
|
| Growth protocol |
NSG mice of 8-9 weeks were sublethally irradiated (250cGy). 24 hours after 2000000 cells MLL-7 PDX were transplanted via tail vein injection.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Sample preparation for scRNA-sequencing was performed by thawing viably frozen bone marrow cells, which were stained with Draq7 (BioLegend), hCD45 and hCD19, together with a PerCP/Cyanine5.5 anti-mouse CD45. Approximately 20.000 alive/mCD45neg/hCD45pos/hCD19pos cells were sorted into a PBS (GE Healthcare Life Sciences) + 2% FBS (Thermo Scientific) coated tube and prepared for sequencing using the 10x Genomics platform at the Center for Translational Genomics, Lund University
Chromium NEXT GEM Single Cell 3' Reagent Kits User Guide (v3.1 Chemistry Dual Index)
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| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
scRNAseq after AZD1775 (Wee1i) treatment,Chromium Single Cell 3ʹ Reagent Kits v3
|
| Data processing |
PDX cell scRNA-seq data was processed and aligned with Cell Ranger (version 6.0.1) using GRCh38-2020-A genome as reference.
Cells were filtered with Cell Ranger 6.0.1 following alignment.
Utilization of R package Seurat (v3) enabled further analysis. MLL-7 data was filtered with the following cutoffs: cells had more than 1500 but less than 6000 UMIs and percentage of mitochondrial counts less than 10%
Assembly: GRCh38-2020-A
Supplementary files format and content: barcode annotation: filtered barcode annotation; feature annotation: filtered feature list; count matrix: filtered UMI count matrix
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| |
|
| Submission date |
Dec 07, 2022 |
| Last update date |
May 15, 2023 |
| Contact name |
Merja Heinäniemi |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Eastern Finland
|
| Department |
School of Medicine, Institute of Biomedicine
|
| Lab |
Systems Genomics - Heinäniemi lab
|
| Street address |
Yliopistonranta 1E
|
| City |
Kuopio |
| ZIP/Postal code |
70211 |
| Country |
Finland |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE220114 |
Mechanistic study of Wee1 kinase inhibition with AZD1775 exposes drug targetable vulnerabilities in acute B-lymphoblastic leukemia |
| GSE220476 |
Mechanistic study of Wee1 kinase inhibition with AZD1775 exposes drug targetable vulnerabilities in acute B-lymphoblastic leukemia [scRNA-seq PDX MLL-7] |
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