Introduction of Bcor frameshift mutations with CRISPR/Cas9 lead to pro-B1 ALL in NP23 HSPCs. (A) Survival curve for recipient mice transplanted with WT/EV, WT/Bcor sgRNA1 (Bcor), NP23/EV, and NP23/Bcor lin⁻ BM or FL cells. Survival curve represents pooled results from 6 experiments and includes recipients of transduced cells (defined as GFP PCR⁺ and >1% CD45.2⁺ at 1 month posttransplantation). P value is indicated (log-rank Mantel-Cox test), *P < .05; **P < .01. (B) Complete blood count (CBC). (C) Flow cytometry of BM and spleen from mice WT, T259, A1929, A1942, A1961, A1959, and A1964 stained with B220, CD19, and CD45.2 antibodies. Red arrow indicates residual normal B cells; blue line outlines the malignant pro-B1 population. Expression of B220 and CD19 were analyzed by gating CD45.2⁺ live cells in the A1929, A1942, and A1961 mice, whereas the expression of B220 and CD19 were analyzed by gating live cells in the T259, A1959, and A1964 mice. (D) May-Grunwald Giemsa–stained peripheral blood (left) and BM cytospin (right) from mouse T259. Scale bars, 50 μm. (E) Igh DJ and VDJ rearrangement assay for the malignant cells of T259. *Clonal Igh rearrangement. HGB, hemoglobin; PLT, platelet; SP, spleen; WBC, white blood cell.