CDK6 is required for reconstitution of the hematopoietic system. (A) Schematic representation of experimental design. Cdk6^+/+/Ly5.1⁺ and Cdk6^−/−/Ly5.2⁺ BM cells were transplanted in ratios of 1:9, 1:1, and 9:1 into lethally irradiated (9 Gy) wild-type recipient mice. Long-term BM reconstitution was analyzed 16 weeks after transplantation (n = 5 per genotype; Cdk6^+/+ and Cdk6^−/− groups were compared using Student t test). (B) Ly5.1⁺/Ly5.2⁺ compositions were analyzed in total BM leukocytes (n = 5 per genotype; ****P < .0001). The leukocytic population included lymphocytes and myeloid cells, but excluded debris and erythrocytes, as determined by forward scatter/side scatter blots. (C) LSK cells were analyzed for Ly5.1⁺/Ly5.2⁺ composition in each transplantation setting (n = 5 per genotype; ****P < .0001). (D) Contributions of Ly5.1⁺ and Ly5.2⁺ cells in long-term HSCs (LT-HSCs), short-term HSCs (ST-HSCs), and MPP (n = 5 per genotype; ****P < .0001, *P < .05). (E) Cdk6^+/+ and Cdk6^−/− BM were infected with empty dsRed⁺ or GFP⁺ retrovirus. Equal numbers of Cdk6^+/+ dsRed⁺ LSKs and Cdk6^−/− GFP⁺ LSKs (100 000/mouse) were injected in a 1:1 ratio into lethally irradiated recipient animals in a competitive setting (LSK-depleted carrier BM was added and a total of 3 × 10⁶ cells/mouse were injected). After 18 hours, mice BM was analyzed for dsRed⁺ and GFP⁺ LSKs (left panel, n = 6). In a noncompetitive setting, equal numbers of either Cdk6^+/+/Ly5.2⁺ or Cdk6^−/−/Ly5.2⁺ BM cells were injected into lethally irradiated Ly5.1⁺ mice (1 × 10⁶ cells/mouse). After 18 hours, BM was analyzed for the presence of donor-derived Ly5.2⁺ LSKs (right panel, n = 4 per genotype).