Glyphosate is a widely used herbicide that inhibits the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS)-encoding aroA gene in the shikimate pathway. The discovery and cloning of the aroA gene with high resistance is central to breeding a transgenic glyphosate-resistant plant. A novel aroA(pantoea) gene from Pantoea was previously isolated and cloned. 'I he aroA(pantoea) enzyme was defined as a new class I EPSPS with glyphosate resistance. The aroA(pantoea) gene was introduced into tobacco through Agrobacteriummediated transformation. he transgenic tobacco plants were confirmed by PCR, RT-PCR, and Southern blot. The analysis of glyphosate resistance also showed that the transgenic tobacco plants could survive at 15 mM glyphosate; the glyphosate resistance level of the transgenic plants is higher than the agricultural application level recommended by most manufacturers. Overall, this study shows that aroA(pantoea) can be used as a candidate gene for the development of genetically modified crops.