Background and Aims Studies have shown that abnormal expressions long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) and the interaction between them play important roles in the occurrence and development of malignant tumors. This study was conducted to investigate the expression characteristics of lncRNA MIR31HG and miR-101 in differentiated thyroid cancer tissues of patients, as well as their relationship with clinicopathologic features and prognosis.Methods The expression levels of lncRNA MIR31HG and miR-101 in the surgical specimens of tumor tissue and adjacent non-tumor tissue form 92 patients with differentiated thyroid cancer were detected using qRT-PCR. The clinical and follow-up data of the patients were collected, and the relationship between the expressions of these two molecules and clinicopathologic factors was analyzed. Kaplan-Meier analysis was used to analyze patient survival rates, and the Cox proportional hazards regression model was used to analyze factors for poor prognosis in patients.Results The relative expression level of lncRNA MIR31HG was significantly higher while the relative expression level of miR-101 was significantly lower in differentiated thyroid cancer tissue than those in adjacent non-tumor tissue (both P<0.01). The relative expression levels of lncRNA MIR31HG and miR-101 were significantly associated with clinical stage, differentiation degree, and lymph node metastasis of patients (all P<0.05). Patients with high expression of lncRNA MIR31HG had lower cumulative survival rates than those with its low expression (84.7% vs. 94.6%, χ²=7.032, P=0.016), and patients with low expression of miR-101 had lower cumulative survival rates than those with its high expression (78.3% vs. 95.6%, χ²=8.482, P=0.004). Clinical stage Ⅲ-Ⅳ, high differentiation degree, lncRNA MIR31HG relative expression level >1.5, and miR-101 relative expression level <0.5 were identified as risk factors for poor prognosis in differentiated thyroid cancer (all P<0.05).Conclusion Upregulation of lncRNA MIR31HG and downregulation of miR-101 in DTC tissues are risk factors for poor prognosis in differentiated thyroid cancer, suggesting that they could serve as prognostic markers for differentiated thyroid cancer.