Live-animal imaging of native hematopoietic stem and progenitor cells

As the biology of hematopoietic stem cells (HSCs) has been predominantly studied under transplantation conditions, it has been challenging to study dynamic HSC behaviors in their native niche given under steady state conditions. Here, we describe the generation of a double knock-in fluorescent reporter that restricts the reporter labeling exclusively to the LT-HSC compartment. Single cell RNAseq comparing previously published LSK signatures (GEO: GSE90742) to our sorted fluorescently labeled cells (sorted without the use of any other cell surface markers) demonstrates that the fluorescently marked cells in our unique mouse model are exclusively found in the LT-HSC compartment in terms of their overal RNA content. This confirms that the generated mice label LT-HSCs in a highly specific manner. Overall design: Single-cell mRNA sequencing of GFP+ long-term hematopoietic stem cells isolated from Mds1-GFP/+ Flt3-Cre mouse bone marrow. As GFP cells represent a very rare fraction of the BM, LT-HSC cells from wild type mice were sorted for the SLAM markers and mixed with the GFP LT-HSCs to ensure high quality of derived libraries. Sample GFP LT-HSCs represents 46 single cells in total after processing while the rest of cells are SLAM LT-HSCs (not used for analysis).