Tregs did not affect the TCR breadth of Tcons after HCT. (A) Schematic representation of the experimental pipeline. On day 0, Balb/c or C57Bl/6 recipient mice were lethally irradiated and received transplantation with 5 × 10⁶ CD45.1⁺ Thy 1.2⁺ TCD-BMs with or without 1 × 10⁶ Foxp3^GFP Tregs from C57Bl6 donors. On day 2, 1 × 10⁶ CD45.1⁺ Thy1.1⁺ Tcons from C57Bl/6 donors were injected. GFP⁺ donor Tregs and CD45.1⁺ Thy1.1⁺ CD4 and CD8 donor Tcons before transplantation (day 0 of 2) and isolated on day 8 were used for sequencing analysis. (B-G) Clonality of the TCRA and TCRB repertoire in CD4 (B) and CD8 (E) Tcons recovered at day 8 after HCT in syngeneic recipients (green box and symbols), allogeneic recipients (blue box and symbols), and allogeneic recipients receiving Tregs (red box and symbols). Representative example of overlap of the TCRA and TCRB repertoire in CD4 (C) and CD8 (F) Tcons before transplantation and at day 8 after HCT (left panels). Scatter plots (C,F) represent clone frequencies before and after HCT and the number of unique clones (dot size). Clones that are only observed at 1 time point are colored in light gray, whereas overlapping clones are colored in dark gray. Repertoire overlap in CD4 (D) and CD8 (G) Tcons recovered at day 8 after HCT and before injection quantified using the Jaccard index of similarity. Groups were compared using a nonparametric Mann–Whitney U test and P values are shown.