Hi, I'm trying to calculate a ratematrix for a RNA aligment (U instead of T) in order to use it as a ratematrix in Profidst (a phylogenetic program which takes into account both the primary sequence and the secondary structure of the RNA, in my case rRNA). The sequence-structure aligment has been made in 4SALE (a java app) and saved as one-letter encoded (using a 12 letters alphabet, "a","c","d","e","g","h","i","k","l","n","q","r" instead of the conventional nucleotide codes). My intention is to calculate the ratematrix (which is a 12x12 matrix) for this special aligment with ape and phangorn, however I've repeatdly fail to do it. The aligment is in a phyDat object containing 30 sequences and 12 states (by using "user-defined" character states consisting on the 12 letters indicated above). I follow the steps described in the phangorn-specials vignette but the ratematrix (under a GTR substitution model) is not calculated. May the problem be that phangorn only accepts "a","g","c" and "t" as valid states for calculating the matrix? And in case phangorn could calculate the matrix, how could I do it? I have a very basic knowledge of R so please I would greatly appreciate (if possible) a step-by-step explanation. Thanks very much for the help. -- View this message in context: http://r.789695.n4.nabble.com/phangorn-and-calculation-of-a-rate-matrix-tp4550495p4550495.html Sent from the R help mailing list archive at Nabble.com.