苹果酸脱氢酶基因(mdh)与肌肉的生长密切相关，可以通过影响骨骼肌的能量代谢来调节肌纤维的生长及类型转化，为探索mdh基因在团头鲂中的功能及作用，实验利用CRISPR/Cas9技术对团头鲂mdh进行敲除，并对敲除突变体的表型及基因表达变化进行了研究。结果显示，CRISPR/Cas9可成功应用于团头鲂mdh基因的敲除，与对照组相比，敲除突变体个体大小明显小于对照组，生长性状数据显示，其体重、体长均发生显著变化。qPCR结果显示，敲除突变体中mdh的表达水平显著低于对照组，同时，在敲除突变体中，生肌决定因子及肌纤维类型因子基因 MyoG 、 MyHCⅡa 的表达水平随着mdh表达水平的下降显著降低，而MyHCⅡb的表达未发生显著变化。研究表明，CRISPR/Cas9可用于团头鲂基因的高效、快速编辑，也证明mdh基因敲除后可能会抑制团头鲂的生长，在团头鲂的生长过程中起着促进作用。这一研究结果为团头鲂的生长性状研究提供了一定的理论基础，有利于mdh基因在团头鲂分子育种中的合理应用。

Malate dehydrogenase gene (mdh) is closely associated with muscle growth, regulating muscle fiber growth and type transformation by influencing skeletal muscle energy metabolism. This study aimed to investigate the function and role of mdh in Megalobrama amblycephala by using CRISPR/Cas9 to knock-out the mdh and examining the phenotype and gene expression in the resulting mutants. The findings demonstrated that CRISPR/Cas9 can effectively knock-out mdh in M. amblycephala, with mutants exhibiting significant phenotypic differences from the control group. Mutants were notably smaller, with reduced body weight, length, height and width compared to controls. qPCR results showed that the expression level of mdh in the mutant individuals was significantly lower than that in the control group. In addition, the expression of MyoG, MyHCⅡa decreased significantly in mutants with reduced mdh, while MyHCⅡb showed no significant change. All the results suggest that CRISPR/Cas9 can be used for efficient and rapid gene editing. It also proved that the mdh gene may plays a promoting role in the growth of muscle in M. amblycephala, and the knock-out of mdh will inhibit the growth. The results of this study provide a certain theoretical value for the study of growth traits, which is conducive to the rational application of mdh in molecular breeding of M. amblycephala.

S 965.119

国家自然科学基金(31272633，31201760，31572220)；上海高校知识服务平台(ZF1206)；上海地方高校能力建设规划(18050501900)