Delivery methods for CRISPR/Cas9 gene editing in crustaceans.

Area of Science:

• * Crustacean genomics and gene editing technologies.
• * Molecular biology and genetic engineering.

Background:

• * CRISPR/Cas9 technology enables precise genomic editing.
• * Efficient delivery of CRISPR/Cas9 components is crucial for its application in crustaceans.
• * Embryonic microinjection is the primary method but presents technical difficulties.

Purpose of the Study:

• * To provide an up-to-date overview of CRISPR/Cas9 delivery methods in crustaceans.
• * To offer a technical guide for embryonic microinjection using examples from three species.
• * To introduce and discuss novel delivery techniques for crustacean gene editing.

Main Methods:

• * Review of existing literature on CRISPR/Cas9 delivery in crustaceans.
• * Detailed technical guide for embryonic microinjection in *Daphnia*, *Parhyale hawaiensis*, and *Exopalaemon carinicauda*.
• * Exploration of Receptor-Mediated Ovary Transduction of Cargo (ReMOT Control) and electroporation as alternative methods.

Main Results:

• * Embryonic microinjection is a widely used but technically demanding method for CRISPR/Cas9 delivery in crustaceans.
• * Specific challenges and protocols for microinjection in *Daphnia*, *P. hawaiensis*, and *E. carinicauda* are presented.
• * ReMOT Control and electroporation show potential as alternative, possibly more efficient, delivery systems.

Conclusions:

• * Advancements in CRISPR/Cas9 delivery are essential for expanding genomic editing applications in crustaceans.
• * Embryonic microinjection remains a key technique, with improved protocols enhancing its utility.
• * Emerging methods like ReMOT Control and electroporation offer promising avenues for future research and genetic manipulation in diverse crustacean species.