Error log file

You should check the error log file prefix.errorlog. Typical errors include:

seqname contains non-ACGTU characters. Omitting this sequence.
Aborting: sequence 3, unknown nucleotide 'S', at 2345.
Warning: sequence 3, ambiguous nt 'S', at 2345.
$\rightarrow$ You can set skip = 0 to allow standard ambiguous nucleotide codes.

Fitting number of mutations (87) outside that allowed by given t range. Sequence pair 4.
Iteration on t failed to converge in maxiter iterations. Sequence pair 5.
Fitting number of mutations (987) outside that allowed by given V range. Sequence pair 2.
Iteration on V failed to converge in maxiter iterations. Sequence pair 2.
$\rightarrow$ You may need to change the parameters tttmin, tttmax, tVfit, maxiter, Vmin, Vmax. Alternatively you may have a bad alignment - perhaps one of your sequences is too divergent from the others. (Note that the Fitting number of mutations outside that allowed by given V range will always occur for the nc model since, for the nc model, all nucleotides are assumed non-coding so the $V$ parameter has no effect.)

Warning: seqname.orfs has overlapping same-frame CDSs.
$\rightarrow$ In this case, regions of the second CDS that overlap the first, in the same frame, are ignored.

Warning: gap within coding region not a multiple three nt; sequence pair 3, sequence 1, sequence coord 1280, alignment coords 1296.
$\rightarrow$ These error messages are quite common if you have extra CDSs in some sequences but not in others, or if corresponding CDSs are longer in some sequences than in others. In this case there may be local problems with the estimated expected number of mutations, as codons will be picked from the wrong read-frame in sequence 2. Consider discarding more divergent sequences, supplying *.orfs files for all sequences, or using the range1--range2 parameters to remove the problematic region.

More serious errors include:

Aborting: ORF 3 in seqname.orfs, endpoints outside sequence range.
$\rightarrow$ Check the *.orfs files. Note that these should be in the corresponding sequence coordinates, not alignment coordinates.

Warning: number of nucleotides in ORF 2 in seqname.orfs not a multiple of 3.
$\rightarrow$ The programme will continue to work, but you should check your *.orfs files. Note that for frameshifts, disjoint CDSs and circular genomes, you should use the GENBANK join(2307..3212,1..1623) format in the *.orfs files.