Hello again, I just ran eprimer3 on a multiple FASTA file (using published genome sequences), and noticed a couple of messages:
"Error: Unrecognized base in input sequence" Additionally, for two of the sequences there were no primer pairs (just some blank lines instead). These appear to correspond to two of the sequences in my input which had IUPAC ambiguous characters in the sequence (e.g. R, W, Y, N). The eprimer3 documentation does say explicitly that for some input files such characters are converted into N (options -mispriminglibraryfile and -mishyblibraryfile) . What is supposed to happen in a sequence in the main input file has such characters? I would expect to still get back a candidate set of primers (even if they do not cover the regions with ambiguous letters). As an experiment I added an N character to the end of an unambiguous sequence, and eprimer3 seemed happy. So, as a work around I've simply replaced all the ambiguous characters (like R, W and Y) with N, and it seems to work. Maybe eprimer3 could do this for me, or at least have this limitation mentioned in the documentation? Thanks, Peter C. _______________________________________________ EMBOSS mailing list [email protected] http://lists.open-bio.org/mailman/listinfo/emboss
