MSG: Got a sequence without letters. Could not guess alphabet

[allebiotech]

Hi, I've run Roary on 17 closely related strains but when I try to run the Roary script (pan_genome_core_alignment) to include an outgroup into the core alignment, a warning comes out:

MSG: Got a sequence without letters. Could not guess alphabet .

I've tried to add this: -alphabet=>'dna' in the MergeMultifastaAlignments.pm but nothing changed (maybe I did it wrong? I'm not an expert user).

This issue was labelled as "bug" in 2016. Are there any new feedback?

My summary statistics file says:

Core genes (99% <= strains <= 100%) 1759
Soft core genes (95% <= strains < 99%) 0
Shell genes (15% <= strains < 95%) 2106
Cloud genes (0% <= strains < 15%) 3282
Total genes (0% <= strains <= 100%) 7147

Thanks in advance!

[andrewjpage]

Hi, You need to include the outgroup along with the other files when you run roary. Andrew

…

On Wed, 25 Jul 2018, 14:34 allebiotech, ***@***.***> wrote: Hi, I've run Roary on 17 closely related strains but when I try to run the Roary script (pan_genome_core_alignment) to include an outgroup into the core alignment, a warning comes out: MSG: Got a sequence without letters. Could not guess alphabet . I've tried to add this: -alphabet=>'dna' in the MergeMultifastaAlignments.pm but nothing changed (maybe I did it wrong? I'm not an expert user). This issue was labelled as "bug" in 2016. Are there any new feedback? My summary statistics file says: Core genes (99% <= strains <= 100%) 1759 Soft core genes (95% <= strains < 99%) 0 Shell genes (15% <= strains < 95%) 2106 Cloud genes (0% <= strains < 15%) 3282 Total genes (0% <= strains <= 100%) 7147 Thanks in advance! — You are receiving this because you are subscribed to this thread. Reply to this email directly, view it on GitHub <#410>, or mute the thread <https://github.com/notifications/unsubscribe-auth/AABeV1TrH3UBv_Gcu-9Z8cl_tAA_PIO_ks5uKHPpgaJpZM4VgGAF> .

[Guanguan8]

-------------------- WARNING ---------------------
MSG: Seq [GKOOBEJB_00026]: Terminator codon inside CDS!
--------------------- WARNING ---------------------
MSG: Seq [GKOOBEJB_00027]: Terminator codon inside CDS!
This happens when I annotate genomes by prokka, and I try to run the Roary script ，a same warning comes out:

MSG: Got a sequence without letters. Could not guess alphabet .
Are there any new feedback?

[allebiotech]

Thanks a lot @andrewjpage, it works! 👍

[andrewjpage]

@Guanguan8 Its an issue with Prokka: tseemann/prokka#203

[Guanguan8]

@andrewjpage，
But I'm looking at Mycoplasma's which may use a different translation table，How to solve it？
Thanks in advance!

[vidhya123979]

@andrewjpage sir ,still i have issues for creating pan genome alignment file ,but gff files were correct but it showing

2024/11/06 16:23:06 Running command: /home/dayholf/miniconda3/envs/pangenome/bin/FastTree -fastest -nt accessory_binary_genes.fa > accessory_binary_genes.fa.newick
FastTree Version 2.1.11 Double precision (No SSE3)
Alignment: accessory_binary_genes.fa
Nucleotide distances: Jukes-Cantor Joins: balanced Support: SH-like 1000
Search: Fastest+2nd +NNI +SPR (2 rounds range 10) +ML-NNI opt-each=1
TopHits: 1.00*sqrtN close=default refresh=0.50
ML Model: Jukes-Cantor, CAT approximation with 20 rate categories
Initial topology in 0.02 seconds
Refining topology: 19 rounds ME-NNIs, 2 rounds ME-SPRs, 9 rounds ML-NNIs
Total branch-length 1.543 after 0.37 sec9, 1 of 24 splits
ML-NNI round 1: LogLk = -10688.093 NNIs 2 max delta 2.05 Time 0.54
Switched to using 20 rate categories (CAT approximation)1 of 20
Rate categories were divided by 0.693 so that average rate = 1.0
CAT-based log-likelihoods may not be comparable across runs
Use -gamma for approximate but comparable Gamma(20) log-likelihoods
ML-NNI round 2: LogLk = -10550.112 NNIs 1 max delta 0.00 Time 0.62
Turning off heuristics for final round of ML NNIs (converged)
ML-NNI round 3: LogLk = -10549.999 NNIs 0 max delta 0.00 Time 0.73 (final)
Optimize all lengths: LogLk = -10549.998 Time 0.76
Total time: 0.93 seconds Unique: 26/28 Bad splits: 0/23
Aligning each cluster
Use of uninitialized value in require at (eval 899) line 1.
2024/11/06 16:26:59 Running command: pan_genome_core_alignment -cd 99
2024/11/06 16:26:59 pan_genome_core_alignment -cd 99

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet

--------------------- WARNING ---------------------
MSG: Got a sequence without letters. Could not guess alphabet