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Differential Rates of Evolution
Aside from varying in size, the protein-coding genes in the mt genome vary in their rates of molecular evolution. Among the four genes that meet the 900bp size threshold, COI and cyt b evolve much more slowly than ND4 or ND5. This rate variation has one important practical implication- it is impossible to design universal primers for ND4 and ND5, effectively ruling out their use as the basis for a comprehensive DNA-identification system. By contrast, because the other two genes evolve more slowly, it has been possible to design primers for them with broad taxonomic utility.

Aside from determining the ease with which the DNA needed for sequencing can be recovered, rates of molecular evolution influence the information content of the resultant sequences. Obviously a gene that showed no sequence changes across life would be useless in a taxonomic context. Conversely a gene with extremely high rates of evolution might distinguish closely allied taxa, but could fail to discriminate more distantly related forms because of secondary convergences. In the case of protein-coding genes, information content can be obtained by inspection of both nucleotide sequences and amino acid arrays in the gene products. Because amino acid composition is so conserved, analyses at this level are most useful for identifying higher taxonomic categories (phyla, classes, orders), while studies at the nucleotide level are useful in making species-level identifications.

COI Versus Cyt b
Based on their large sizes and slow rates of molecular evolution, COI and cyt b are the MT genes best suited to serve as the basis for a DNA- identification system. COI does have two important advantages over cyt b, both linked to its slower rate of molecular evolution. Firstly, the universal primers for this gene are very robust, enabling the recovery of its 5' end from most animal species. Furthermore, COI has a greater taxonomic signal range than cyt b. Both genes do show a high incidence of base substitutions at third position nucleotides, allowing the discrimination of closely allied species. However, COI provides better resolution of deeper taxonomic affinities because its amino acid sequence changes more slowly than that of cyt b.