Extended Data Fig. 2: Oncogenic mutations induce neoplastic growth in mini-colons.

a, Time-course brightfield and fluorescence images of non-induced healthy colon cells grown as conventional organoids and mini-colons. Absence of fluorescence signal indicates absence of oncogenic recombination. Scale bars, 200 μm (organoids) and 75 μm (mini-colons). b, Immunofluorescence images showing the expression of Fabp1 (left, green), and Sox9 (right, magenta) in healthy mini-colons cultured for 7 days. Scale bar, 100 μm. c, Fluorescence image (left) showing the presence of mutated cells 36 h after the blue–light-mediated induction of a mini-colon. The segmentation of each mutated cell is shown (right). Scale bar, 120 μm. d, Evolution of the number of mutated cells in an inducible mini-colon after blue–light-mediated activation. Each dot represents a measurement every 15 min. The 2^nd order smoothing of the data is shown. e, Time-course quantitation of cell shedding (total protein content) into the lumen of OptoCre and control mini-colons after blue-light induced oncogenic recombination. *P = 0.0156; **P = 0.0033; ***P = 0.0003 (24 h), <0.0001 (96 h) (two-way ANOVA and Sidak’s multiple comparisons test; n = 3 mini-colons for each condition). Each point represents one mini-colon. Data represent mean ± SEM. f, Low- (left) and high-magnification (right) immunofluorescence images showing the presence of CD44 (magenta) and nuclei (blue) in a tumour-bearing mini-colon. White and grey arrowheads indicate early and advanced tumorigenic events, respectively. Scale bars, 120 μm (left) and 50 μm (right).

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