Use of CRISPR/Cas9 technology efficiently targetted goat myostatin through zygotes microinjection resulting in double-muscled phenotype in goats.

Myostatin gene (MSTN) can inhibit the proliferation of myoblast, which in turn promotes muscle growth and inhibits adipocyte differentiation in livestock. MSTN mutation may lead to muscle hypertrophy or double-muscled (DM) phenotype. MSTN mutation animal, such as sheep, dog, and rabbit have been generated through CRISPR/Cas9 technology. However, goats with promising MSTN mutation have not been generated. We designed two sgRNAs loci targetting exon3 of MSTN gene to destroy the MSTN cysteines knots. We got seven goats from seven recipients, in which six were MSTN knocked-out (KO) goats, with a mutation rate of 85.7%. Destroyed cysteine knots caused MSTN structure inactivation. The average body weight gain (BWG) per day of MSTN KO goats was significantly higher than that of wild-type (WT) goats. MSTN KO goats showed abnormal sugar, fat, and protein metabolism compared with wild-type controls (MSTN(+/+)). Inheritance of mutations was observed in offspring of MSTN KO goats by PCR analysis.