The aim of this study was to detect the expression levels of Dicer1, Drosha, DGCR8, and Ago2 messenger ribonucleic acids (mRNAs) in patients with primary gastrointestinal diffuse large B-cell lymphoma (PGI-DLBCL) and determine their associations with clinical parameters and prognostic significance. The mRNA level expressions of Dicer1, Drosha, DGCR8, and Ago2 were detected by real-time quantitative polymerase chain reaction. Immunohistochemical staining of CD10, BCL6, and MUM1 was performed using EnVision™ system. The clinicopathologic features and follow-up data were analyzed using Kaplan-Meier estimator. The results show that the expression of Dicer1 (P=0.001), Drosha (P=0.01), DGCR8 (P=0.02), and Ago2 (P=0.002) mRNAs in cancer tissues of patients with PGI-DLBCL was significantly lower than those in normal tissues of healthy controls. Among the expression of CD10, BCL6, and MUM1, 27.4% (17/62) of the patients belonged to the germinal center B-cell (GCB) subtype and 72.6% (45/62) belonged to the non-GCB subtype. Dicer1 expression was significantly decreased in the non-GCB subgroup (P=0.02) and in the high International Prognostic Index (3-5 score) subgroup (P=0.03). Kaplan-Meier analysis showed that the low-Dicer1 subgroup had a shorter overall survival (P=0.02) and shorter progression-free survival (P=0.015) than the high-Dicer1 subgroup. Multivariate analysis identified Dicer1 as an independent prognostic factor in PGI-DLBCL. In Conclusion, Dicer1, Drosha, DGCR8, and Ago2 play key roles in the pathogenesis of PGI-DLBCL. Dicer1, an independent prognostic factor for predicting shortened survival of patients with PGI-DLBCL, can be used as a biomarker to guide the prognosis.