Lactoferricin B, the antimicrobially active N-terminal peptide derived from acid pepsin hydrolysis of bovine lactoferrin, has broad spectrum antimicrobial activities. We expressed the lactoferricin gene in tobacco plants to evaluate its antimicrobial activity. e coding region for the chimeric peptide gene of the signal peptide from tobacco pathogenrelated protein (PR-1) and bovine lactoferricin was synthesized using the recombinant polymerase chain reaction (PCR) method. Transgenic tobacco plants expressing the lactoferricin gene were developed using the Agrobacterium-mediated transformation method. e lactoferricin B gene was integrated into the tobacco genome and its transcription was detected by PCR, Southern blot analysis, and reverse transcription PCR (RT-PCR), respectively. e transgenic tobacco plants were challenged with the pathogenic bacteria Pseudomonas syringae pv. tabaci and Botrytis cinerea. At 30 or 28 days postinoculation, the transgenic plants were still green and continued to grow, whereas the control plants were infected with bacterial or fungal pathogens from the roots to the tips, resulting in death of the plants. In conclusion, transgenic tobacco plants that overexpressed the lactoferricin gene, linked to the signal peptide of tobacco PR-1 protein under the control of a high expression constitutive promoter, showed enhanced resistance to bacterial (P. syringae pv. tabaci) and fungal (B. cinerea) diseases.