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. 2022 Mar 15;8:24. doi: 10.1038/s41531-022-00289-9

Fig. 1. Breeding paradigm and immune phenotype characterization of double mutant chimeras.

Fig. 1

a Breeding diagram and BMT strategy used to generate double mutant chimeras, (WT)GFP ->Rag1/R1441G. b Flow cytometry confirmed that similar to Rag1−/− mice, non-transplanted double mutant mice lack both B- and T-lymphocytes regardless of R1441G transgene expression. Note that immune-competent R1441G mice used as a control exhibit normal proportions of adaptive immune cells compared to WT. Data are mean ± SEM, n = 20 for WT and R1441G groups; n = 3–4 for Rag1−/− and double mutant groups. c Representative flow cytometry plot from blood of double mutant chimera 8 weeks after BMT demonstrating significant repopulation with GFP + BM cells. d qPCR analysis showed the lack of human lrrk2 mRNA expression in T- and B-cells sorted from BM-transplanted double mutant chimera’s (WT(GFP) -> Rag1/R1441G). Cells were sorted based on GFP+/CD4 + (CD4-PerCP-CY5.5), GFP+/CD8 + (CD8-PE-CY7) and GFP+/CD19 + (CD19-APC-CY7) fluorescent intensity. Sorted GFP- cells from double mutant chimeras (innate immune cells) were used as a positive control for human lrrk2 mRNA expression. mRNA expression was normalized to mouse β-actin. Data are mean ± SEM, n = 4. e, f Similar to Rag1 chimeras, serum cytokines, and antibodies are back to control R1441G level in BM-transplanted double mutant chimeras. Data are mean ± SEM, n = 5–10 for each group. ND not detectable.