Disruption of mstn Gene by CRISPR/Cas9 in Large Yellow Croaker (Larimichthys crocea).

The large yellow croaker (Larimichthys crocea) plays an economically vital role in the marine aquaculture in China. Suffering from infection of bacteria and protozoon, effect of extreme weather and stress from high-density farming, genome editing is thought to be an important tool applied to L. croea for enhancing commercial traits such as growth rate, disease resistance, and nutrition component. In this study, we identified two mstn genes in L. croea and investigated the different phylogenetic clades, gene structures, and conserved syntenic relationships. To obtain fast-growing large yellow croaker, we specially selected two validated targets for mstnb knockout, which was homologous to mammalian myostatin gene (MSTN) and downregulated skeletal muscle growth and development. Five significant mutation types were generated in two mosaic mutants by transferring specific CRISPR/Cas9 RNPs (ribonucleoprotein) into the one-cell fertilized embryos based on CRISPR/Cas9 technology. Subsequently, we also elucidated the obstacles and possible measures to improve the success rate of inducing modified large yellow croaker. Our results would provide valuable method and reference for facilitating genome editing programs of the large yellow croaker in the future.