Motivated by the successful optimizations of MAFFT and MUSCLE parameters, we searched for optimal gap-costs of CLUSTALW [27,28]. We varied gap-open (go) and gap-extension (ge) penalties from 7.5 to 22.5 and from 3.33 to 9.99, respectively (default values of CLUSTALW for RNA/DNA sequences are go = 15.0 and ge = 6.66, respectively). Ranks derived by Friedman tests are averaged over all alignment sets, i. e. consisting of 2, 3, 5, 7, 10, and 15 sequences. Table 2 summarizes the results. Alignments created with higher gap-open penalties score significantly better. A combination of go = 22.5 and ge = 0.83 is optimal for the tested parameter range. It should be noted that this performance gain results mainly from a better SCI, whereas the SPS' remains almost the same.