Hello,

the MACSE_V2 toolkit provides several tools to deal with nuceotide coding sequences. The alignSequences subprogram of MACSE allows building reliable codon alignments even in the presence of frameshifts of stop codons (especially useful for dN/dS analysis and pseudogene analysis). Morevover, this subprogam can handle the fact that different sequences use different genetic codes. MACSE also includes a subprogram specifically designed to replace stop codons (and frameshift codons) from an alignment. This subprogam (exportAlignment) allows to specify the codon (three letters of your choice) that will replace the stop codons. You can even provide two different codons for replacing stops appearing within the sequence (unexpected unless in pseudogenes) and stop codons appearing at the end of the sequences. While there is several options (e.g. to specify the output file name and the genetic code to use) the basic usage is quite straightforward:

java -jar macse.jar -prog exportAlignment -align align.fasta -codonForFinalStop --- -codonForInternalStop NNN

To ease the alignment of coding nucleotide sequences, we also provide ready to use alignment pipelines (provided as singularity container), which include optional filtering steps. These pipelines output the (filtered) nucleotide alignment, the corresponding (filtered) amino acid ones and the detail of the filtering steps (if some filtering steps were selected).

Hi,

Few time ago i got the same problem. I solved using a perl script available here (. Since is needed to feed with both nuclotides and amino acids i have used t-coffee to translate.

This have worked fine for me. I have done in linux, for windows you may neeed to write a .bat file to it easily. Take a look on bat files tutorial for syntax if you are not familiar with that. I think that will work.

You can use fasta format as sequence file for PAML no need of .pml format.

Regards,

Joao

Pal2Nal will generate a codon alignment without stop codons, given a MSA of proteins and the corresponding DNA sequences. If the input is a pairwise alignment, I believe it will calculate dN/dS ratios (using PAML) for you automatically. Otherwise, you can just input your alignments to PAML to calculate dN/dS. Pal2Nal is written in Perl, so it should work on your Win7 machine (I don't know about PAML though, unless there is a Windows version available).