Our present study included the following highlights: (1) Dicer knockout cells were developed using the CRISPR/Cas9 system; (2) The efficacy of tetra-looped DsiRNA was compared with linear DsiRNAs in colorectal carcinoma cell lines; (3) The gene silencing activity of various DsiRNAs disappeared in Dicer knockout cells; (4) The nicked tetra-looped DsiRNA required functional Dicer protein for silencing activity. The gene discussed is DICER1; the disease is colorectal carcinoma.