1 gt rosa 26 sortm1 eyfp cos j rosa eyfp reporter mice  (Jackson Laboratory)

Journal: Cell reports

Article Title: Mds1 CreERT2 , an inducible Cre allele specific to adult-repopulating hematopoietic stem cells

doi: 10.1016/j.celrep.2021.109562

Figure Lengend Snippet: (A) A DNA cassette encoding the TAM-inducible Cre-ER fusion protein was introduced into the Mecom locus in embryonic stem (ES) cells via homologous recombination. (B) ES cell DNA was screened by Southern blotting, cutting the genomic DNA with EcoRV and blotting with 5′ and 3′ probes situated outside of the recombination cassette. Clones B3 and B5 show the expected sizes of 15.1 Kb and 7.6 kb, with 5′ and 3′ probes, respectively. Mice were created from ES cell clone B5, which were backcrossed onto a C57Bl6 background and crossed with mice bearing the Rosa26 LSL-YFP allele. (C–E) These mice were treated with TAM, and the peripheral blood was analyzed by flow cytometry at 30 days and bone marrow at days 4, 10, and 15 for YFP + cells, which progressively increased. (D) Quantitation of percentage of bone-marrow YFP-positive cells following the first TAM treatment on the days indicated. Average ± SEM, n = 3. (E) Representative scatterplot (day 30) showing the appearance of YFP + blood cells following TAM treatment. (F and G) X-gal staining of fetal liver in E13.5 mouse embryos, after a single treatment with TAM at E7.5 (F) or E9.5 (G). Eosin counter-stain; bar denotes 10 μm. (H) Flow cytometric analysis of E11.5 AGM, yolk sac, and blood for pre-HSC (kit + CD31 + VECadherin + ) versus EMP (kit + CD41 + CD16/32 + ) as indicated. For each analysis, the percentage of total cells that were YFP + is charted as average ± SEM, n = 5. See and , for the gating strategy. (I) Flow cytometric analysis of E11.5 blood and liver for lineage-positive cell subsets (EryP Ter119 + blood, EryD Ter119 + FSC hi to distinguish from circulating EryP in the liver) as indicated. For each analysis, the percentage of total cells that were YFP + is charted as average ± SEM, n = 5. See and for the gating strategy. (J) Flow cytometric analysis of E12.5 fetal liver and blood for cell subsets as indicated gated as in (I). For each analysis, the percentage of total cells that were YFP + is charted as average ± SEM, n = 10. (K) Analysis of mice competitively transplanted with 10 6 E12.5 liver cells from TAM-treated (at E9.5) Mds1 CreERT2/WT , Rosa26 LSL-YFP/LSL-YFP mice and 10 5 unlabeled adult bone-marrow cells. Flow cytometric analysis of circulating YFP + and YFP-negative granulocyte/monocyte cells (CD11b + ), B cells (CD19 + ), and T cells (CD3e + ) at 12 weeks post-transplant. See for gating strategy. Average ± SEM, n = 3.

Article Snippet: Rosa26-YFP reporter mice ( ) Gt(ROSA)26Sor tm1(EYFP)Cos/J , Stock No: 006148, abbreviated Rosa26 YFP ) and C57BL/6 were obtained from Jackson Labs.

Techniques: Homologous Recombination, Southern Blot, Clone Assay, Flow Cytometry, Quantitation Assay, Staining

Journal: Cell reports

Article Title: Mds1 CreERT2 , an inducible Cre allele specific to adult-repopulating hematopoietic stem cells

doi: 10.1016/j.celrep.2021.109562

Figure Lengend Snippet: (A) Flow cytometric analysis of the proportion of YFP + granulocyte/monocyte cells (GR-1 + ), B cells (B220 + ), and T cells (CD3ε + ) in the circulation of 1-year-old Mds1 CreERT2 Rosa26 LSL-YFP mice after TAM treatment at E8.5 (empty circles, n = 4) or at E9.5 (solid circles, n = 2). (B) Flow cytometric analysis of bone marrow of the same mice as in (A) demonstrating a high percentage of YFP + progenitor cells in LSK subsets, as well as of lineage-restricted lymphoid (CD127 + ), myeloid (CD16/32 + ), and erythroid/megakaryocyte (CD16/32 − ) LK subsets. See for gating strategy. Total marrow CD45 + YFP + cells shown in (C). (C) Percentage of YFP + F4/80 + tissue-resident macrophages in these mice. See for detailed gating strategy. In all panels, the average is plotted.

Article Snippet: Rosa26-YFP reporter mice ( ) Gt(ROSA)26Sor tm1(EYFP)Cos/J , Stock No: 006148, abbreviated Rosa26 YFP ) and C57BL/6 were obtained from Jackson Labs.

Techniques: