Abstract
A comprehensive experimental system for Japanese anchovy, a promising candidate model organism for marine teleosts, was established. Through the design of a rearing/spawning facility that controls the photoperiod and water temperature, one-cell eggs were continuously obtained shortly after spawning throughout the rearing period. The stages of eggs are indispensable for microinjection experiments, and we developed an efficient and robust microinjection system for the Japanese anchovy. Embryos injected with GFP mRNA showed strong whole-body GFP fluorescence and the survival rates of injected- and non-injected embryos were not significantly different, 87.5% (28 in 32 embryos) and 90.0% (45 in 50 embryos), respectively. We verified that the Tol2 transposon system, which mediates gene transfer in vertebrates, worked efficiently in the Japanese anchovy using the transient transgenesis protocol, with GFP or DsRed as the reporter gene. Finally, we confirmed that genome-editing technologies, namely Transcription Activator-Like Effector Nucleases (TALEN) and Clustered Regulatory Interspaced Short Palindromic Repeats (CRISPR)/Cas9, were applicable to the Japanese anchovy. In practice, specific gene-disrupted fishes were generated in the F1 generation. These results demonstrated the establishment of a basic, yet comprehensive, experimental system, which could be employed to undertake experiments using the Japanese anchovy as a model organism for marine teleost fish.