Navigating Eukaryotic Genome Annotation Pipelines: A Route Map to Using BRAKER, Galba, and TSEBRA

Annotating the structure of protein-coding genes represents a major challenge in the analysis of eukaryotic genomes. This task sets the groundwork for subsequent genomic studies aimed at understanding the functions of individual genes. BRAKER and Galba are two fully automated and containerized pipelines designed to perform accurate genome annotation. BRAKER integrates the GeneMark-ETP and AUGUSTUS gene finders, employing the TSEBRA combiner to attain high sensitivity and precision. BRAKER is adept at handling genomes of any size, provided that it has access to both transcript expression sequencing data and an extensive protein database from the target clade. In particular, BRAKER demonstrates enhanced accuracy even with only one type of these extrinsic evidence sources, although it should be noted that accuracy diminishes for larger genomes under these conditions. In contrast, Galba adopts a distinct methodology utilizing the outcomes of direct protein-to-genome spliced alignments with miniprot to generate training genes and evidence for gene prediction in AUGUSTUS. Galba has superior accuracy in large genomes if protein sequences are the only source of evidence. This chapter provides practical guidelines for employing both pipelines in the annotation of eukaryotic genomes, with a focus on insect genomes.