Mus musculus
Protocols: To isolate stromal and endothelial cells from fetal bone, 9 fetuses were collected from a pregnant C57BL/6J mouse euthanized by cervical dislocation at day 17 of gestation (E17). Fetal long bones were dissected and cleaned using a dissection microscope. Collagenased bone fraction and bone marrow were obtained as previously described (Zambetti et al., 2016) and cell suspensions were stained using the following antibodies: anti-CD45.2 (clone 104, 1:100), anti-Ter119 (clone TER-119, 1:50), anti-CD51 (clone RMV-7, 1:50), anti-CD144 (clone BV13, 1:200), anti-Sca1 (clone D7, 1:100) all from Biolegend and anti-CD31 (Clone MEC 13.3, 1:100) from BD Biosciences. To discriminate between live and dead cells, staining with the viability dye 7-AAD (1:100) was performed. Endothelial cell enriched cell populations (7AAD- CD45- Ter119- CD31+) and stromal cells (7AAD- CD45- Ter119- CD31- Sca1+ CD51+) were FACS sorted. Sorted populations from bone marrow and bone fraction were pooled together and counted. An input of 17,000 cells was used for droplet based single cell RNA sequencing. Single cells were encapsulated for cDNA synthesis and barcoded using the Chromium Single Cell 3' Reagent Kit v3 (10X Genomics). Library construction was performed according to the manufacturerвЂ™s recommendations.
ENA first public
2021-01-21
ENA last update
2021-01-11
External Id
SAMEA7778285
INSDC center alias
Dept. of Hematology Erasmus MC
INSDC center name
Dept. of Hematology Erasmus MC
INSDC first public
2021-01-21T04:08:49Z
INSDC last update
2021-01-11T18:07:27Z
INSDC status
public
Submitter Id
E-MTAB-9994:plugging_E17
broker name
ArrayExpress
cell type
bone marrow cell
common name
house mouse
developmental stage
embryonic day 17.5
genotype
wild type genotype
immunophenotype
mixed 7AAD- CD45- Ter119- CD31+ and 7AAD- CD45- Ter119- CD31- Sca1+ CD51+
individual
pool of 9 fetuses
organism part
bone marrow
sample name
E-MTAB-9994:plugging_E17
strain
C57BL/6J