CRISPR/Cas9-knockout of the myostatin gene in common carp

The goal of the study is to develop a method for knocking out the common carp two paralogous copies of the myostatin gene using the CRISPR/Cas9 system with delivery of the editing complex into zygote by microinjection.

Materials and Methods: fertilization of common carp eggs was carried out in vitro, sgRNA was synthesized by PCR with overlapping oligonucleotides and subsequent transcription, the editing complex consisting of sgRNA and the Cas9 endonuclease was delivered to the carp zygote by microinjection before the first cleavage. The region contained the editing site was amplified by nested PCR, editing events were detected by Sanger sequencing and confirmed by NGS.

Novelty: For the first time in Russia, a knockout system for the common carp myostatin gene has been developed, allowing both paralogous copies of the gene to be modified simultaneously.

Result: 12 F0 common carp individuals with knockout of the myostatin gene were obtained.

Practical significance: A methodological approach has been tested that allows for the rapid and controlled production of common carp with programmed mutations. Myostatin gene-deficient fish exhibit increased muscle growth due to hypertrophy and hyperplasia of muscle fibers, resulting in improved fish fillet quality. Fast-growing fish populations are more suitable for aquaculture than populations with standard growth rates.

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