In vivo lymphoid lineage development from coRAG2-targeted healthy donor-derived HSPCs. (A) Schematic of engraftment protocol of human HSPCs into NSG immunodeficient mice and secondary analysis. (B) Percent human cell chimerism (CD45⁺ HLA A-B-C⁺ double-positive cells) in bone marrow (BM) and spleen (SP) of mice 22 weeks posttransplant with CB (intra-hepatic [IH] transplant [Tx]) or (C) PB-derived HSPCs (intra-femoral [IF] transplant [Tx]) edited with RAG2-sgRNA guide 3 (RNP, light grey circles in panel B) or targeted with coRAG2 cassette (GTed, yellow and red circles in panel B, dark grey and red circles in panel C). Each dot represents an individual mouse; panel B: WT (n = 10), RNP (n = 5), GTed (n = 11); panel C: WT (n = 5), GTed-i53 (n = 5), GTed+i53 (n = 6). Bars, median (D) percent coRAG2-GTed by ddPCR before (-Tx) and after (+Tx) transplant of CB and (E) PB-derived CD34⁺ HSPCs into NSG mice and FACS sort from BM, as marked. Bars, mean ± s.e.m.; -i53, without p53 inhibitor; +i53, with p53 inhibitor; stats. One-way ANOVA, nonparametric test, Kruskal-Wallis test, Dunn’s multiple comparisons test.