Based on transcriptome analysis and RT-PCR techniques, a PR10 gene was isolated from P. notoginseng root and named PnPR10. Bioinformatics and phylogenetic tree analyses revealed that the open reading frame (ORF) of PnPR10 was 465 bp in length, encoding 154 amino acids and containing one typical conserved domain of the pathogenesis-related protein Bet v I family, with high similarity to the gene from P. ginseng. Using the genome walking method, a 458 bp promoter upstream of the PnPR10 transcription start site was isolated from P. notoginseng root genomic DNA by hiTAIL-PCR. The PnPR10 promoter contained multiple cis-acting regulatory elements. The CGTCA-motif and TGACG-motif, which have been shown to play important roles in response to MeJA, were identified. The expression of PnPR10 can be induced by typical root rot disease pathogens (F. solani and C. destructans). The purified recombinant PnPR10 protein could clearly inhibit the growth of two pathogens. The results demonstrate that the PnPR10 gene is involved in the defense response against root rot disease pathogens. This experiment lays a theoretical foundation for revealing the mechanism of resistance in P.notoginseng.