Gastric cancer is one of the most common carcinomas in China. microRNAs, a type of non-coding RNA, are important specific regulators and are involved in numerous bioprocesses of an organism. microRNA-21 (miR-21) has been identified as the most suitable choice for further investigation because it is overexpressed in nearly all solid tumors; furthermore, it has been demonstrated that miR-21 is involved in the genesis and progression of human cancer. It has been reported that PTEN, an important tumour suppressor, is regulated by multiple miRNAs. Thus, in this study we focused on the expression and significance of miR-21 in gastric cancer tissues, and the role of miR-21 in the biological behaviour and the expression of PTEN in gastric cancer cells. Real-time PCR was used to detect miR-21 expression in gastric cancer tissues, the adjacent normal tissues, and the gastric cell lines. The gastric cancer cell line BGC-823 was transfected with pre-miR-21/miR-21 inhibitor to overexpress/downregulate miR-21. The influence of miR-21 on the biological behaviour of gastric cancer cells was evaluated using the CCK-8 kit, FCMs, the scratch healing assay and the transwell test. Western blotting and the Luciferase Reporter Assay were used to evaluate the change of PTEN expression after lowered expression of miR-21 in gastric cancer cell lines. Real-time PCR analysis indicated that miR-21 exhibited higher expression in gastric cancer tissues compared to the adjacent non-tumor tissues. miR-21 expression was significantly associated with the degree of differentiation of the tumour tissues (P=0.004), as well as local invasion and lymph node metastasis (P<0.01). After transfection, pre-miR21 BGC-823 cells grew faster than the negative and control groups (P<0.01). The reduction in miR-21 expression demonstrated a remarkable effect on the biological behaviour of gastric cancer cells (P<0.05); the pre-miR-21-transfected cells healed more quickly compared to the control cells in the scratch healing assay, whereas the transwell test indicated that cell migration in vitro was notably inhibited with the downregulation of miR-21 (P<0.05). The western blot results and Luciferase Reporter Assay demonstrated that PTEN expression was remarkably increased after miR-21 inhibition (P<0.05). microRNA-21 expression was upregulated in gastric carcinoma tissues and was significantly associated with the degree of differentiation of tumour tissues, local invasion and lymph node metastasis. Overexpression of miR-21 promoted BGC-823 cell growth, invasion and cell migration in vitro, whereas downregulation of miR-21 exhibited a stronger inhibitory effect on the biological behaviour of gastric cancer cells; additionally, miR-21 inhibition may upregulate the PTEN expression level, which indicates that PTEN may be a target gene for gastric cancer initiation and development.