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Acting as a typical harpin protein, Hpa1 of <i>Xanthomonas oryzae</i> pv. <i>oryzae</i> is one of the pathogenic factors in hosts and can elicit hypersensitive responses (HR) in non-hosts. To further explain the underlying mechanisms of its induced resistance, we studied the function of the most stable and shortest three heptads in the N-terminal coiled-coil domain of Hpa1, named N21_Hpa1. Proteins isolated from <i>N21</i>-transgenic tobacco elicited HR in Xanthi tobacco, which was consistent with the results using N21 and full-length Hpa1 proteins expressed in <i>Escherichia coli</i>. N21-expressing tobacco plants showed enhanced resistance to tobacco mosaic virus (TMV) and <i>Pectobacterium carotovora</i> subsp. <i>carotovora</i> (<i>Pcc</i>). Spraying of a synthesized N21 peptide solution delayed the disease symptoms caused by <i>Botrytis cinerea</i> and <i>Monilinia fructicola</i> and promoted the growth and drought tolerance of plants. Further analysis indicated that N21 upregulated the expression of multiple plant defense-related genes, such as genes mediated by salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) signaling, and genes related to reactive oxygen species (ROS) biosynthesis. Further, the bioavailability of N21 peptide was better than that of full-length Hpa1_Xoo. Our studies support the broad application prospects of N21 peptide as a promising succedaneum to biopesticide Messenger or Illite or other biological pharmaceutical products, and provide a basis for further development of biopesticides using proteins with similar structures.