To understand the spatio-temporal expression pattern of miR-133a-5p and the effect of short-term fasting on circadian rhythm of its expression in Siniperca chuatsi, real-time fluorescence quantitative PCR was used to detect the miR-133a-5p expression in different tissues and embryonic developmental stages and its circadian rhythm after short-term fasting. The results showed that the expression of miR-133a-5p was highest in slow-twitch muscles, higher in fast-twitch muscles and myocardium, but lower in other tissues. The expression level of miR-133a-5p was high at the two-cell stage during embryonic develop-ment, and gradually decreased with the development of the embryo. It maintained at a low level from the late blastula stage to the hatching larvae with no significant difference between each stage. The expression of miR-133a-5p showed no obvious circadian rhythm in fast-twitch muscles of normally fed S. chuatsi, but a circadian rhythm appeared after 5 days of fasting, with high expression during the day and low at night. Taken together, miR-133a-5p was specifically highly expressed in muscle related tissues, suggesting that it may play an important role in the muscular tissue. Its high expression during embryonic development revealed that the target genes regulated by miR-133a-5p may inhibit early embryonic development of S. chuatsi. The rhythmic expression of miR-133a-5p after fasting indicated that miR-133a-5p may be involved in the adaptive regulation of S. chuatsi muscles in response to fasting stress by regulating biological rhythm.