This CAREER project will investigate one of biology’s most fundamental questions: how entirely new genes with novel biological functions arise and help organisms adapt to challenging environments. The project uses fish antifreeze proteins, molecules that allow some fishes to survive in icy seawater, as a window into how evolution repeatedly solves the same problem through different genetic routes. By revealing how genomes generate new functions, the work will advance basic knowledge in evolutionary biology and genomics, with broader relevance to biotechnology and data-intensive biology. The project also serves the national interest by strengthening STEM education and workforce development. It will bring hands-on evolution outreach to K–12 students and families in Arkansas through annual museum events, integrate project-generated multi-omic datasets into classroom research experiences for more than 100 college students, and provide mentoring and research training for students and postdoctoral scholars. Together, these activities will expand public scientific literacy, broaden participation in genomics, and prepare future scientists to analyze complex biological data. The research will use type I antifreeze proteins (AFPI), which occur in four distantly related fish lineages and appear to have evolved independently despite their similar protein sequences and functions. The project will combine comparative genomics, transcriptomics, epigenomics, and phylogenetic analyses to determine how these genes originated, how they acquired regulatory elements and became integrated into gene networks, and how they were maintained, diversified, or lost under different environmental conditions. Specifically, it will analyze strategically selected AFPI-bearing species and close AFPI-lacking relatives to identify precursor sequences and mechanisms of gene birth; map gene expression, open chromatin, histone-marked regulatory regions, and promoter-enhancer interactions to reconstruct