I have trouble working with roary tool. I used the Prokka tool to annotate nineteen genomes and then applied the roary tool to analyze all gff files for pangenome analysis. But there is always error. I tried using only three test genomes but still did not work.
I am using Prokka and Roary in https://usegalaxy.eu/ . There are several errors in the execution of the pipeline.
-Prokka does not output results in .gff3 format, depite the fact that there is an option in Prokka parameters for gff version 3 output.
-Roary will not work with the output of Prokka and will output an error.
-When renaming the .gff files to .gff3 extention, Roary runs successfully, however with default settings, it rarely finds core genes, and all of the genes identified shift to cloud or shell genes.
2026 Update! All of these tools have had major updates, so please try again and any new issues, please open a new ticket so we can investigate! Also, be sure to check out our latest updates to our suite of tutorials! Genome Annotation / Tutorial List