The suprachiasmatic nucleus (SCN) is the master clock that orchestrates circadian clocks across the body to synchronize with and anticipate the earth’s light/dark cycles. Although post-transcriptional regulators called microRNAs have been implicated in physiological SCN function, how the absence of the entire mature miRNome impacts SCN output has not yet been explored. Here, we have generated an SCN-specific Dicer knockout mouse model by crossing Syt10Cre mice with Dicerflox mice to study behavioral consequences of miRNA depletion in the SCN. We show that loss of all mature miRNAs in the SCN shortens the circadian period length by ~40 minutes at the tissue level, and by ~50 minutes at the locomotor activity level. Knockout animals also showed arrythmicity or ultradian locomotor activities with no light masking under constant light, a condition which usually caused lengthening of the circadian period length and reduced activities, i.e. light masking, in nocturnal animals. Moreover, induction of Dicer knockout by Cre injection into the SCN of adult Dicerflox mice eventually resulted in loss of behavioral rhythms. Finally, we show suggestive evidence that SCN desynchronization might be one mechanism underlying the behavioral phenotypes of SCN-specific Dicer knockout animals.