traits within a short period of time. Improved breeds are essential for increasing aquaculture production (Gjedrem et al., 2012). CRISPR/Cas9 technology has great promise in aquaculture for increasing the production as conventional selective breeding usually requires substantial time to achieve desired genetic and phenotypic change compared to gene editing.
Increased muscle growth can be attained by manipulating growth regulating genes using genome editing. Leptin is a hormone secreted by peripheral adipose tissues which regulates appetite, homeostasis and growth in animals, but its functional role has not been studied in blue catfish (Ictalurus furcatus). In this study, CRISPR/Cas9 gene editing was employed to generate leptin knock out (leptin-KO) blue catfish to evaluate growth performance. Two guide RNAs (sgRNAs) targeting exon 1 and exon II of blue catfish leptin gene were designed and co-injected with Cas9 protein into fertilized blue catfish eggs with a ratio of 1:1 gRNA to Cas9. A total of 300 eggs were injected. Hatching percentage in injected and non-injected groups (wild-type control) were 27.6% and 66.6% respectively. Successful mutation was confirmed by amplifying the target gene followed by T7 endonuclease assay. Leptin-KO fish and their wild type controls were reared under identical conditions. Growth parameters evaluated were body weight, length, specific growth rate and percentage weight gain. Mutation efficiency was 100%. After 600 days of culture in aquaria, the leptin-KO had higher (P<0.05) mean body weight (247.40g ± 21.06) compared to the wild-type control (213.09g ± 9.42). The survival rate of leptin-KO and wild-type controls were not different (P>0.05), 53.01% and 55%, respectively.
Leptin-KO blue catfish grew 16.1% faster than wild-type full-sibling controls demonstrating a functional role of leptin in somatic growth of blue catfish. This study demonstrates the feasibility of CRISPR/Cas 9 gene editing in the blue catfish for improved growth.