MiR-21, the only microRNA (miRNA) found to be overexpressed in any type of solid tumor, its guide stand, miR-21-5p, has been studied a lot in colorectal cancer (CRC); however, few researchers focused on its passenger strand, miR-21-3p. In our study, based on The Cancer Genome Atlas (TCGA) data, we found that there were more varieties and quantities of miR-21-3p isoforms in microsatellite instability (MSI)-type CRC. We further examined the role of miR-21-3p by in vitro and in vivo studies. MiR-21-3p may be an oncogene in CRC by promoting cellular mobility through epithelial-mesenchymal transition. However, different isoforms, especially miR-21-3p 0 | 2, may be a favorable prognostic marker for CRC survival, probably due to increased complementary effect of miR-21-5p and/or target genes. Further study investigating the underlying mechanism of miRNA isoforms is needed.