We, together with the Flow Cytometry Unit, FACs purified hematopoietic stem cells from aged (21 month old) and young (3 month old) C57BL/6 male mice. We isolated two sub-populations of HSCs, which we have previously defined as myeloid-biased HSCs and lineage balanced HSCs. Cells from 15 young and 5 old mice (pooled 3 young mice for adequate cell numbers) for a total of 20 samples- 5 young CD150high , 5 CD150low, 5 old CD150high , and 5 old CD150low. We encountered a non-trivial amount of organic contamination in our initial RNA purifications. We are currently optimizing our RNA extraction protocols as any phenol contamination in the RNA isolation leads to significant alterations in the RNA-seq results. Towards addressing the effect of CR on the epigenetic landscapes, we have continued to calorically restricted (CR) C57Bl/6 male mice acquired from the NIA aged CR rodent colony to 24 months of age. Bone marrow analysis and HSC purification from these cells has been performed.
