We used the Brachyury positive CRC cell line, SW480, to derive colonospheres (potentially enriched for cancer progenitor cells), and carried out an extreme limiting dilution assay (ELDA) [34] to determine the ability of single CRC cells to proliferate and form spheres in the presence of Brachyury or under conditions of siRNA-induced Brachyury-depletion. Here, TBX1 is linked to colorectal carcinoma.