Fig. 1: Translocations during SpCas9-mediated multiplex gene editing in human T cells.

a Schematics for assessing chromosomal translocations during multiplex gene editing by SpCas9 RNPs in human T cells. Chr represents Chromosome. b Editing efficiency for TRAC, TRBC, and PDCD1 in human T cells at 3-, 7-, and 14-days post-transfection detected by PEM-seq. Mean ± SD from three biological replicates, represented by “circle”, “triangle”, and “square”, respectively. Paired two-tailed t-test, *p < 0.05 and **p < 0.01. c Circos plot indicating translocations among TRAC, TRBC, PDCD1, and the TRAC off-target in human T cells at 3-days post-transfection detected by PEM-seq. Chromosomes are shown with centromere to telomere in the clockwise direction. The averages of three replicates are shown on each line, and the arrows indicate the orientation of bait to prey. d Percentages for translocations among TRAC, TRBC, and PDCD1 detected by PEM-seq in human T cells at 3-, 7-, and 14-days post-transfection. The translocations were categorized as bait-prey, e.g., TRAC-TRBC represented the translocations between TRAC and TRBC cloned with a bait primer located at TRAC. Mean ± SD from three biological replicates. e Percentages for general translocations in human T cells cloned from the indicated loci at 3-, 7-, and 14-days post-transfection detected by PEM-seq. Mean ± SD from three biological replicates. Mean values are marked above each data point. f Gene annotation for total translocations from TRAC, TRBC, and PDCD1 in human T cells at 3-days post-transfection using the KEGG feature of Enrichr (maayanlab.cloud/Enrichr/). The horizontal axis indicates the gene numbers in the indicated pathway. g Percentages for the indicated translocations induced by SpCas9 and indicated variants detected by PEM-seq in human T cells cloned from TRAC at 3-days post-transfection, N = 1. Source data are provided as a Source Data file.