Mutational profiling was performed by SNaPshot analysis (left panels) and confirmed by Sanger sequencing (right panels). Nucleic acid extracted from each MCC archival specimen was run in parallel with a genomic DNA control, as indicated. The arrows point to the mutant alleles: PIK3CA Q546K (c.1636C>A) in tumor MCC-12 (A), PIK3CA E545K (c.1633G>A) in MCC-13 (B), and PIK3CA E542K (c.1624G>A) for MCC-20 (C). The assayed loci for each panel are as follows: (A) 1.PIK3CA 3140, 2.CTNNB1 101, 3.BRAF 1798, 4.NRAS 37, 5.PIK3CA 1636, 6.APC 4348 and 7.APC 3340; (B) 1.KRAS 34, 2.EGFR 2235_49del F, 3.EGFR 2369, 4.NRAS 181, 5.PIK3CA 1633, 6.CTNNB1 94 and 7.CTNNB1 121; and (C) 1.EGFR 2236_50del F, 2.EGFR 2573, 3.CTNNB1 133, 4.PIK3CA 1624 and 5.NRAS 35.