Direct PCR sequencing on genetic material containing allelic mixtures results in sequences containing ambiguous nucleotides. Because codons exhibiting allelic mixtures present evidence of evolutionary pressure, it is important to include this information in the assessment of codon synonymy. We developed a program, Synonymous-Nonsynonymous Mutation Rates between Sequences Containing Ambiguous Nucleotides (Syn-SCAN), that calculates synonymous and non-synonymous substitution rates using a model that includes allelic mixtures.