Figure 3

Some colorectal cancer cell lines are viable and can be propagated in the absence of β-CATENIN. (a) Schematic representation of the CTNNB1 gene and its protein product β-CATENIN. Exons are depicted as numbered boxes. Exon 7 (red box) was chosen for CRISPR/Cas9-induced deletion using two sgRNAs targeting positions marked by red arrowheads. The location of the start codon (AUG) in exon 2 and functionally important domains of β-CATENIN are indicated and connected to the exons by which they are encoded. (b) Western blot analyses to detect β-CATENIN (β-CAT) expression. Cell lysates were generated from cells with biallelic WT and mutant CTNNB1 genes. α-TUBULIN (αTUB) or GSK3β were used as loading controls. Molecular weights are given in kDa. Representative results from one of three independent biological replicates are shown. Full size/uncropped versions of the Western blot images are shown in Supplementary Fig. S7. (c) Representative micrographs from one of three independent biological replicates showing HCT116, HT29, SW480 and RKO cell clones with biallelic WT and mutant CTNNB1 genes. Images were taken 24 h after seeding the same starting numbers of cells. The scale bars represent 50 µm.