Fig. 3
Evaluation and applications of HiHo-AID2 in other human cancer cell lines. a–f Comparison of degron-GFP tagging efficiencies in coIN with different HDR enhancers in A549 (a, b), HEK293A (c, d), and U2OS (e, f) cells. Representative FACS profile (a, c, e) and graphs depicting the percentage of GFP-positive cells, the percentual change of single-cell GFP intensity compared to control (coIN), and the cell count (× 105 cells/ml) (b, d, f) are shown. Numbers above columns indicate mean values; lines link the same endogenous tagging pairs. N = 6 pairs of tagging plasmids. Statistical comparisons are shown in Additional file 4: Table S6. g Graphs showing the genotyping PCR results for AID clones generated with HiHo-AID2. Clones were generated and identified as indicated in Fig. 2f, g. Total number of clones analyzed is indicated above each column. N.A.: not available due to cell death after selection with blasticidin (S2). h WB analysis of inducible SAC1 degradation in A549 and HEK293A cells. i Graphs showing the PI4P staining intensity upon SAC1 degradation in A549 and HEK293A wild-type and SAC1 AID cells. N = 17 (A549) and 20 (HEK293A) fields. One-way ANOVA, n.s.: non-significant, **** p < 0.001. All statistical comparisons are shown in Additional file 4: Table S6. j Widefield imaging of cell morphological changes upon SAC1 degradation. Scale bar: 50 μM. Representative of 1 (A549) and 2 (HEK293A) clones (h–j). WT: wild-type; pico: 0.5 μM pico_cvxIAA treatment; a.u.: arbitrary unit