Gastric cancer (GC) is one of the most common malignant tumors with high mortality and metastasis rate. Previous studies elucidated that Long non-coding RNA (LncRNA) MEG3 was down-regulated in various tumors and participated in tumor progression. The aim of our study was to investigate the regulating role of MEG3 in cell proliferation and metastasis of GC cells. Our data showed that down-regulated MEG3 was observed in GC tissues and cell lines (MKN74, MKN45, SGC7901, AGS). Overexpressed MEG3 by pcDNA3.1-MEG3 transfection suppressed cell proliferation, migration and invasion of GC cells remarkably. Besides that, the targeting relationship between MEG3 and miR-21 was firstly revealed by bioinformatics prediction. Overexpressed miR-21 was observed in GC tissues and cell lines. Results from luciferase report assay indicated that miR-21 was a target of MEG3 and results from qRT-PCR indicated that the expression of miR-21 was negatively regulated by MEG3. Moreover, overexpressed miR-21 promoted cell proliferation and metastasis. However, pcDNA3.1-MEG3 transfection could counteract the promoting role of miR-21 mimic on GC cell proliferation and metastasis, indicating that MEG3 suppressed proliferation and metastasis of GC cells through inhibiting miR-21 expression. Finally, the mice tumor model experiments showed that overexpressed MEG3 could also inhibit tumor growth and metastasis in vivo through inhibiting miR-21 expression. In summary, our study revealed the regulating role of MEG3/miR-21 axis in GC progression and provided a new potential therapeutic strategy for GC treatment.