PAM matrices were introduced by Margaret Dayhoff in 1978.[7] The calculation of these matrices were based on 1572 observed mutations in the phylogenetic trees of 71 families of closely related proteins. The proteins to be studied were selected on the basis of having high similarity with their predecessors. The protein alignments included were required to display at least 85% identity.[2][8] As a result, it is reasonable to assume that any aligned mismatches were the result of a single mutation event, rather than several at the same location.