Primaclade didn find any primers for my alignment! What should I do now?
Increase the number of allowable degenerate nucleotides in your final primer product and try it again. Also, take a look at your alignment and pay close attention to the consensus line. In the consensus line non-degenerate sites are labeled ACGT whereas degenerate sites use the IUPAC ambiguity codes. In our experience, if sequences in your alignment are greater than 29% divergent then you aren’t likely to find primers that are going to work well across it (see Tests of the Program). I increased the Max Number of Degenerate nucleotides, but still no luck. Try increasing the Alignment Gaps to Skip parameter. Once again, take a close look at your alignment file. One or two sequences with a number of gaps can throw Primaclade off. If you have a sequence that has several insertions that cause gap columns in the alignment, you might want to try deleting that sequence and resubmitting the new alignment. The same applies to a sequence with a large number of spaces in it. Sometimes removing jus
