EDGE: Expanding the functional genetics toolkit to link genes to phenotypes in cichlid fish

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How does the genome encode the information for proper development and function of the body? This important question is technically and ethically difficult to address in humans, and there is a need for additional animal models that can support gaining this foundational understanding. The cichlid fish are an excellent system to use because these species have wide variation in anatomy, physiology, and behavior. Because their genomes have been sequenced recently, correlations between animal features and gene sequences can be determined. However, the causal links between gene sequence and function must be tested directly to gain meaningful understanding. Causal testing of the links requires the development and deployment of new technologies. The work supported by this grant develops the tools to obtain cichlid embryos and modify gene sequences, enabling researchers to understand how specific genes control the form and function of the body. Given that cichlids and humans share most of the same genes, this work should allow new insights into how genes function in health and disease. The work also allows many opportunities for undergraduate researches from diverse backgrounds to participate directly in the research enterprise, and it includes workshops for investigators from other institutions to enable them to adopt the new protocols and tools in their own research programs.The 2000 species of cichlid fish exhibit a stunning variety of phenotypes in morphology, physiology, and behavior. Genome-wide scans have mapped these traits to candidate genetic loci. However, functional genetic manipulations are required for direct tests of gene function, and technology for such experiments has lagged behind. CRISPR/Cas has emerged as a potent gene editing tool, and proof-of-principle experiments have been performed in two cichlid species. The work of this grant involves development of experimental pipelines that enable gene editing experiments in a wider variety of cichlid species. Specifically, hormonal treatment regimens that support the reproductive cycle of cichlids and derivation of fertilized eggs are identified, and protocols that permit the modification of genes in a variety of species, through creation of loss-of-function mutations and the insertion of species-specific sequence variants into other species, are developed. Insights and knowledge obtained through these efforts are shared with the broader community through workshops, websites, and publications. Together, the newly developed approaches will support the study of genetic variation and its effects on form and function.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.