Background: Panax notoginseng (Burk.) F. H. Chen (PN) belonging to the genus Panax of family Araliaceae is widely used in traditional Chinese medicine to treat various diseases. PN taproot, as the most vital organ for the accumulation of bioactive components, presents a variable morphology (oval or long), even within the same environment. However, no related studies have yet explained the molecular mechanism of phenotypic differences. To investigate the cause of differences in the taproot phenotype, de novo and comparative transcriptomic analysis on PN taproot was performed. Results: A total of 133,730,886 and 114,761,595 paired-end clean reads were obtained based on high-throughput sequencing from oval and long taproot samples, respectively. 121,955 unigenes with contig N50 = 1,774 bp were generated by using the de novo assembly transcriptome. The annotation results showed 38,313 BLASTX hits and 24,820 BLASTP hits. KEGG analysis identified 52 genes encoding class III peroxidase (PRX) family members, 13 genes encoding L-Ascorbate peroxidase (APX) family members, and 99 genes encoding a series of mitogen-activated protein kinase (MAPK) family members. Differentially expressed genes analysis indicated substantial upregulation of genes encoding APX3 and PRX45 enzymes, which are related to reactive oxygen species metabolism, and the MPK3 enzyme, which is related to cell proliferation and plant root development, in long taproots compared with that in oval taproots. Furthermore, the determination results of real-time quantitative PCR, enzyme activity, and H2O2 content verified transcriptomic analysis results. Conclusion: These results collectively demonstrate that reactive oxygen species (ROS) metabolism and the MPK3 enzyme may play vital roles in regulating the taproot phenotype of PN. This study provides further insights into the genetic mechanisms of phenotypic differences in other species of the genus Panax.