SLAC

[Kaparepen]

I don't know what I do when the mistake is: The alignment must include at least 3 unique sequences for selection methods to work. Could you help me please?

[spond]

Dear @Kaparepen,

This error message tells me that HyPhy loaded your alignment, discarded all identical sequences (retaining a single copy for all identical sequences), and ended up with 2 or fewer sequences. You cannot do a proper comparative analysis on ≤3 sequences with SLAC. All you can do, really, is estimate pairwise dN/dS if you have 2 sequences.

What was your input alignment?

Best, Sergei

[Kaparepen]

Thanks for the response. I have 8 sequences for the same protein but each sequence is from different bacterial isolates, and I need to analyses the present SNPs in these sequences. The first sequence is from the reference strain. How could I do the dN / dS analysis?

King regards, Karen

[spond]

Dear @Kaparepen,

Sorry, this information is insufficient for me to provide meaningful help. If you could attach your alignment here, that would be best. Barring that, please copy and paste the output from SLAC when you run your data through.

Best, Sergei

[Kaparepen]

Thanks for the response. Output SLAC : The alignment must include at least 3 unique sequences for #selection methods to work Input Input peptidase M4.txt

NZ_CP011849.2:c3148091-3146307/1-1782 Piscirickettsia salmonis LF-89 = ATCC VR-1361 chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCGATTACTGCATCTACAGCACTGTTATCATCCCATGCA TTAAGCACGGTAAATTCATATAATAAGCCTGTTTCCTCTTCTAATATTGAAGGTTTTGCGATTGAAGGTGCA GGAGGTAGCTCAAATACGATGCTGTCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGAATGGGGATACTTACGTACGTTACCAGCAAAAATATGAGGGTATTCCTGTAATTGGT AAGCAGGTAGTGGTAAAACAGCCTAAAGCTGTTACTGGCTTTGCAGCAACGAGCCGATCGGCGTCGAGAGCT ACAGCTACTCGAATATCTTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTAAGCGCTGGTGATGCC ATGGCGTTTGCAAAACAGCAGTTTGAGCAGAGCTATAGTGGTACACAAGTTGCTGATGGTTCTAATTCTGTT AAGGCAACAAAAGAGATTCGTATTGTAGATAATAAAGCTCGGCTTTATTATCGAGTGACGTTTAATGCTAGT AATACAGCTGGTGGTAAGCCATATAGTATGGTTTATATTATTGCTGCGAACGGTGGGGCTAAGCCTGTAGTG CTTAAGCATTGGGACAATATTCAAAATTATGAAGATACTGGCCCTGGTGGAAATGAGAAAACGGTAAAGCAT GGTCCAACAGGTGTTGAATTTTTTTATGGAGAAAATAATTTACCAGCATTGAATGTGAGTGAGAATAACGGC AGTTGCACAATGGACAATGGAGATGTTCGGCTGGTTGATGTGCAGAATCAAGAGGATCACTCTTGGGATAGT GATTACAATACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCAATCAATGGTGCGTATTCA CCCACAGATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCT TTGCAAGAAAATGGTGAGCCAATGCAGTTAATTATGCGTGTACATTATGGTACTGATTATGACAATGCATTT TGGGATGGACAGACCATGTCATTTGGCGATGGTAGTAGCTTTTATCCACTTGTATCTTTGGATGTCGCAGGT CATGAAGTTAGCCATGGATTTACTGAGCAGCATTCGGGTTTAGAATATAGTGATCAATCAGGTTCATTAAAT GAAGCCTTTTCTGACATGGCAGGCCAGGCAGTGCGAGCTTATTTATTAAGTACAAACTCTGACTTATACAAG CAACTGTATTTTAATCAAGATGAAGTCACTTGGGGGATTGGTGAGACTATCATGAAAGGTGATAATACAGAT ACGGCTTTGCGTTATATGGATCAGCCTTCTAAAGATCAAGATGAAAATGGTGTTTCGGCAGATTGTTTAGAT AAGGATTTAGCAGGATCAGGGTGTATTATATCATATGATGATGTTGTCACCGCAGCAAAAAAACTCCCACTT CGCTATCAGCAAAGTTATATCGTTCACCATGGAAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAA CAGGTGGGTATTAAAGAAGCCTTTAAAGTGATGAAGGATGCGAATGCTACACGTTGGACTTCAGGTTCAGAT TTTGCAGATGCTGCATGTGGTGTGCTTCAGGCAGCTCATGCTGATGGTGTGGGTTCTGACTCAATGATTAAA GAAGTTTTTAATCAAGTGGGGGTTGCTATAGTAGATGAGGACTGCTCTACTAAG NZ_CP038893.1:36700-38432/1-1733 Piscirickettsia salmonis strain Psal-006a chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT------------------------------------------------- NZ_CP038908.1:35453-37185/1-1733 Piscirickettsia salmonis strain Psal-009 chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT------------------------------------------------- NZ_CP038923.1:35443-37175/1-1733 Piscirickettsia salmonis strain Psal-011 chromosome, complete genome Peptidasa M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT------------------------------------------------- NZ_CP038811.1:35443-37175/1-1733 Piscirickettsia salmonis strain Psal-001 chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT------------------------------------------------- CP038891.1:35443-37175/1-1733 Piscirickettsia salmonis strain Psal-005 chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT------------------------------------------------- NZ_CP038972.1:35443-37175/1-1733 Piscirickettsia salmonis strain Psal-069 chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT------------------------------------------------- NZ_CP039040.1:35453-37185/1-1733 Piscirickettsia salmonis strain Psal-072 chromosome, complete genome Peptidase M4 ATGCGAAAAAAAAATCTTGCTATTATTATTTCTGCCATTACTGCATCTACTGCACTATTATCATCCCATGCA TTAAGCACGGTAAATTCATACAATAAGCCTGTTTCCTCTTCTAATATCGAAGGTTTTGCGATTGAAGGTGCA GGAGATAGCTCAAATACAATGCTATCAAGTCGCTCTGCTACAACGACTGATAAAAATACCCTAAGCCAAGTG AGTAGTACGACAATGGATGGCGATACTTACATCCGTTACCAGCAAAAATATGAGGGTATTCCTGTAATCGGT ACACAGGTGGTGGTAAAACAGCCTAAAGCTGTTACTGGTTTTGCAGCAACGAGCCGATCGGCATCGACGGCA ACAGCTACTCGAATATCGTTGGCAAAAGATTTAGATGTTGATTTGGTTGCAACAGTGAGTGCTGATGATGCC AAAGCATTTGCTAAACAGCAATTTGAAAAGGATTATAGTGGTACGCAAGTTGCTGATGACTCTATAAAGTCA ACAAAAGAGATTCGTATTGTAGATAATAAAGCTCAGCTTTATTATCGAGTGACGTTTAATGCTAGCAATACA GCTGGTGGTAAGCCATATCGTATGGTTTATATTATTAATGCCACTGGGACCTCTAAGGCTATGGTGCTGAGT TTCTGGGACAGTATTCCAAATTATAGTGATACTGGGCCTGGTGGAAATGAGAAAACGGTACAGTATGGTCCA ACAGGCGTTAAATTTTTTTATGGGGAAAATAATTTACCAGCCTTGAATGTGAGTGAGGACAACAGCACCTGC ACGATGGACAATGGAGATGTTCGGTTGGTTAATGTAGAGCATCAAGCAGATCATTCTTGGGATAGTGATTAC AACACAACGGCTTATCAATATAGCTGTGGCCACAATCAGGGTGATCCGATCAATGGTGCGTATTCACCTACA GATGATGCTTATTATTTTGGTAGTATGATTATTGATATGTATAAGAATTGGTATGGTGTTGATGCTTTGCAA GAAAATGGCGAGCCAATGCAATTAATTATGCGTGTCCATTATGGTACTGATTATGATAATGCATTTTGGGAT GGGCATACTATGTCATTTGGCGATGGTAGTAGCTTTTATCCTCTTGTATCTCTGGATGTTGCAGGTCATGAA GTGAGCCATGGATTTACTGAGCAGCACTCTGGTTTGGATTATAAATATCAATATGGATCATTAAATGAAGCC TTTTCTGATATGGCAGGTCAAGCGGTGCGGGCTTATTTATTAAGTACAAATCCTGATTTATATAAGCAACTG TACTTTAACCAAGATGAAGTTACTTGGGGTATTGGTGAAACAATTTCTAAAGGGGATGATGAGAGTGATGCC CTACGTTATATGAATAACCCTTCTAAAGATGGAGTCTCGGCAGATTGTGATGATAAGGAATTAGCTGGATCG ACGTGTACTATATCATATGATGAGGTAGTAGCTACCTCAAAAGACTACCCAATTAAAGATCGACAGAGTTAT ATTGTTCATACTGGTAGTGGTGTATTTAATAAGGCGTTTTACTTGTTATCACAACAGGTGGGTATTAAAGAT GCCTTTAGGGTAATGAAGGATGCGAATGTTAAATATTGGGGTAAATACTCAGATTTTTCAGATGCTGCATGT GGTGTGCTAAAAGCTGCTAATGACGACGGTGTGGGTTCTACCTCAATGATTAAAGATGTTTTTAATCAAGTC GGGGT-------------------------------------------------

[spond]

Dear @Kaparepen,

Out of 8 sequences in your alignment, 6 are duplicates (identical copies); Datamonkey.org will automatically prune those out, yielding 2 sequences (and the error). If you run it locally in HyPhy, e.g.

hyphy slac --alignment ~/Downloads/Input.peptidase.M4.txt --tree neighbor-joining

You will see the following message

-------
>[WARNING] This dataset contains 6 duplicate sequences. Identical
sequences do not contribute any information to the analysis and only
slow down computation. Please consider removing duplicate or 'nearly'
duplicate sequences, e.g. using
https://github.com/veg/hyphy-analyses/tree/master/remove-duplicates
prior to running selection analyses
-------

Best, Sergei

[github-actions[bot]]

Stale issue message

[Stradichenko]

Good evening; Like @spond, I'm having this issue; which is strange to me because in my case I'm using 200 sequences, many with different lenghts and nucleotides, from a big variety of species, very visible in MEGA. So is unclear to me what identical refers to. In my case I just want to do a dn/ds (ka/ks) but I'm unclear why is not processing any input.

[stevenweaver]

Dear @Stradichenko,

Just as a sanity check, did you align the sequences first before trying SLAC?

Best, Steven

[Stradichenko]

Hi @stevenweaver, I was double checking... I did an Muscle alignment and the format is .mas (from MEGA). I don't know if this may be involved somehow. The data comes from Ensembl Compara orthologs fasta CDS for Olig2: https://www.ensembl.org/Homo_sapiens/Gene/Compara_Ortholog?db=core;g=ENSG00000205927;r=21:33025935-33029196

Thank you for your attention, Gary.

[spond]

Dear @Stradichenko,

Could you please include your alignment as a text attachment for me to check what's going on?

Best, Sergei

[Stradichenko]

Hi @spond thank you for your quick response; I double-double checked and it was an error on my side while clearing the stop codons in sequences. To solve that I tried the code from this post but I ran with the following error on my file named example.fas in my conda env named env_name:

$ hyphy pre-msa.bf --input example.fas
Error:
Could not read batch file '/home/.../ensembl-data/pre-msa.bf'.
Path stack:
        /home/.../.conda/envs/env_name/lib/hyphy/
        /home/.../ensembl-data/

Check errors.log for execution error details.

EDIT: I used instead the command: hyphy -? > (8) Data File Tools > (3) Convert sequence names to HyPhy valid identifiers if needed and replace stop codons with gaps in codon data if any are present.

And it worked, I'm not sure if this conveys the same result of clearing stop codons as the post's method but, the result file was fortunately accepted by the Datamonkey site.

Best, Gary

[spond]

Dear @Stradichenko,

pre-msa.bf is not a standard analysis, so HyPhy will look for the file in the same directory that you are calling in from (ensembl-data), and then the HyPhy installation directory (/.conda/envs/env_name/lib/hyphy/).

You need to download codon-msa as a part of hyphy-analyses , and then either call HyPhy from the codon-msa directory or do something like

 $hyphy /path/to/pre-msa.bf ...

Best, Sergei