Ethylene-responsive factors (ERFs) are a subfamily of the AP2/ERF transcription factor superfamily in plant, whose members play an important role in regulating gene expression in plant responses to biotic and abiotic stresses. A soybean mRNA sequence encoding an ERF protein was cloned and designated GmERF5 from gene expression profiles of Jilin 32 immature embryo by use of RT-PCR. The GmERF5 consists of an ORF of 714 bp and encodes 237 amino acids (26.09 kDa) with an isoelectric point of 6.85. GmERF5 is grouped with Class IV ERF transcription factors and is related most closely to GhERF2. Real-time quantitative RT-PCR analysis revealed that GmERF5 expressed highly in root. GmERF5 was upregulated when Jilin 32 was treated by drought, high salt, low temperature, ethylene, abscisic acid and methyl jasmonate. GmERF5 protein was localized in the nucleus by subcellular localization assay in onion epidermal cells. The regulation ability of GmERF5 was analyzed in tobacco leaves by transient expression. GmERF5 activation of a reporter gene indicated that GmERF5 had transcription activation activity. GmERF5 may be involved in soybean biotic and abiotic stress responses as a transcription factor.