A ribozyme-mediated, gene “knockdown” strategy for the identification of gene function in zebrafish

The zebrafish system offers many unique opportunities for the study of molecular biology. To date, only random mutagenesis, and not directed gene knockouts, have been demonstrated in this system. To more fully develop the potential of the zebrafish system, an approach to effectively inhibit the expression of any targeted gene in the developing zebrafish embryo has been developed. This approach uses a transient, cytoplasmic, T7 expression system, injected into the fertilized zebrafish egg to rapidly produce high levels of a ribozyme directed against the mRNA encoded by the targeted gene to inhibit its expression. In a demonstration of this strategy, expression of the recessive dominant zebrafish no tail gene was effectively inhibited by using this strategy to yield a phenotype identical to that resulting from a known defective mutation in this same gene. This, ribozyme-mediated, message deletion strategy may have use in determining the function of genetic coding sequences of unknown function.