Targeted transgene integration overcomes variability of position effects in zebrafish

摘要: Zebrafish transgenesis is increasingly popular owing to the optical transparency and external development of embryos, which provide a scalable vertebrate model for in vivo experimentation. The ability express transgenes tightly controlled spatio-temporal pattern an important prerequisite exploitation zebrafish wide range biomedical applications. However, conventional methods are plagued by position effects: regulatory environment genomic integration sites leads variation expression patterns driven engineered cis-regulatory modules. This limitation represents bottleneck when studying precise function modules their subtle variants or various effector proteins be expressed labelling manipulation defined sets cells. Here, we evidence efficient elimination variability effects developing PhiC31 integrase-based targeting method. To detect targeted events, simple phenotype scoring colour change lens larvae used. We compared PhiC31-based Tol2 analysis activity novel conserved enhancer from developmentally regulated neural-specific esrrga gene. Reporter was highly variable among independent lines generated with Tol2, whereas all into single site displayed nearly identical, enhancer-specific reporter brain nuclei. Moreover, demonstrate that modified integrase system can also used detection transient transgenesis. These results power transgene annotation fine transcriptional elements it promises generally desirable tool applications, rely on reproducible zebrafish.