Annotations for: A new phased assembly of the Antarctic spiny plunderfish provides novel insights into the evolution of the notothenioid radiation

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Notothenioids are a well characterised species flock endemic to the Antarctic and an important model group for the study of genome adaptation to extreme cold. We used a new reference assembly and clade-wide comparative genomic analysis to investigate cryonotothenioid evolution and the appearance of novel functionalities linked to cold adaptation. Gene annotation: Gene annotation of Hap1 was performed with BRAKER3. Repetitive elements were soft masked and both protein and transcriptomic data were used as external evidence to train AUGUSTUS and GeneMark-ETP ab-initio gene predictors. Illumina RNAseq reads from muscle, skin, liver, kidney, heart, and brain and from gills (data generated herein) were mapped with HISAT2. The sorted bam files were supplied to BRAKER3 together with the protein database previously used for cleaning the repetitive libraries from host genes. The quality of the annotation was assessed with BUSCO v.5. To obtain a gene annotation for Hap2, we mapped BRAKER-derived genes with Liftoff (-polish -copies) to obtain their lifted coordinates. Functional annotation was performed on the Hap1 predicted proteome with eggNOG-mapper v2 transferring only annotations with experimental evidence and auto-adjusting the taxonomic scope based on the query sequence. Repeat annotation:Repetitive elements were de-novo mined with RepeatModeler v.2.0.5 adding the -LTRStruct extension to improve LTR detection. Potential multicopy host genes included in the repetitive library were removed with ProtExcluder v.1.2 after blasting (Blastx, e-value 1e-10) the repetitive library against a protein database consisting of all vertebrate proteins included in the Swiss-Prot database (The UniProt Consortium, 2025) and the predicted proteomes of five published notothenioid genomes available on NCBI RefSeq: C. gobio, P.georgianus, Gymnodraco acuticeps, Trematomus bernacchii, Eleginops maclovinus. Furthermore, a TE library was made with MCHelper v.1.7.0 with default parameters. Both haplotypes were annotated with RepeatMasker in sensitive mode (-s).