Standard methods for transgenesis in zebrafish depend on random transgene integration into the genome followed by resource-intensive screening and validation. Targeted vector integration into validated genomic loci using phiC31 integrase-basedattP/attBrecombination has transformed mouse andDrosophilatransgenesis. However, while the phiC31 system functions in zebrafish, validated loci carryingattP-based landing or safe harbor sites suitable for universal transgenesis applications in zebrafish have not been established. Here, using CRISPR-Cas9, we converted two well-validated single insertionTol2-based zebrafish transgenes with long-standing genetic stability into twoattPlanding sites, calledphiC31 Integrase Genomic Loci Engineered for Transgenesis(pIGLET). Generating fluorescent reporters,loxP-based Switch lines, CreERT2 drivers, and gene-regulatory variant reporters in thepIGLET14aandpIGLET24blanding site alleles, we document their suitability for transgenesis applications across cell types and developmental stages. For both landing sites, we routinely achieve 25-50% germline transmission of targeted transgene integrations, drastically reducing the number of required animals and necessary resources to generate individual transgenic lines. We document that phiC31 integrase-based transgenesis intopIGLET14aandpIGLET24breproducibly results in representative reporter expression patterns in injected F0 zebrafish embryos suitable for enhancer discovery and qualitative and quantitative comparison of gene-regulatory element variants. Taken together, our new phiC31 integrase-based transgene landing sites establish reproducible, targeted zebrafish transgenesis for numerous applications while greatly reducing the workload of generating new transgenic zebrafish lines.SHORT ABSTRACTTargeted transgenesis into pre-established, “safe harbor,” landing sites remains a missing technique in zebrafish. Here, we establishedphiC31 Integrase Genomic Loci Engineered for Transgenesis (pIGLET) by CRISPR-Cas9-based conversion of two previously validatedTol2transgenes intoattPsites. phiC31-mediated transgenesis into ourpIGLET14aandpIGLET24b attPlanding sites results in 25-50% germline transmission efficiency and quantifiable transgene activity. Our landing sites are suitable for reproducible, routine zebrafish transgenesis for diverse applications.