CAUSA 2.0: accurate and consistent evolutionary analysis of proteins using codon and amino acid unified sequence alignments

Multiple sequence alignment (MSA) is widely used to reveal structural and functional changes leading to genetic differences among species, and to reconstruct evolutionary histories of related genes, proteins and genomes. Traditionally, proteins and their coding sequences (CDSs) are aligned and analyzed separately, but often drastically different conclusions were drawn on a same set of data. Here we present a new alignment strategy, Codon and Amino Acid Unified Sequence Alignment (CAUSA) 2.0, which aligns proteins and their coding sequences simultaneously. CAUSA 2.0 optimizes the alignment of CDSs at both codon and amino acid level efficiently. Theoretical analysis showed that CAUSA 2.0 enhances the entropy information content of MSA. Empirical data analysis demonstrated that CAUSA 2.0 is more accurate and consistent than nucleotide, protein or codon level alignments. CAUSA 2.0 locates in-frame indels more accurately, makes the alignment of coding sequences biologically more significant, and reveals several novel mutation mechanisms that relate to some genetic diseases. CAUSA 2.0 is available in website www.DNAPlusPro.com .