Efficient Gene Knockdown in Adult Zebrafish Retina by Intravitreal Injection

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Abstract
High-throughput sequencing has created a tremendous amount of information about the genes expressed in various cell types and tissues throughout the body. As such, there is a need for a quick and effective method to knock down genes of interest in order to investigate their roles. While there are many approaches for this in mammalian models, there are limited ways to knock down genes of interest in adult zebrafish. Unlike mammals, zebrafish have the natural ability to regenerate their neurons after injury or disease is detected, making them a staple in regenerative studies. Unfortunately, current approaches for gene knockdown in the retina of adult zebrafish are costly and provide a barrier for many scientists. We provide two cost-effective approaches for targeted gene knockdowns in adult zebrafish retinas. We describe this approach through the use of Vivo-morpholinos and lipid-encapsulated siRNAs that target the expression of the proliferating cell nuclear antigen (PCNA) gene in adult zebrafish. We also describe how to collect and process retina samples for downstream immunohistochemistry, imaging, and quantification. Overall, this protocol will provide researchers with a straightforward, cheap, and effective method to perform targeted gene knockdowns in adult zebrafish retinas.

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