Tree peony cultivars are usually classified according to flower characteristics (flower form and flower color) which are commonly affected by environmental influences and developmental levels. Judgment of flower forms may also depend on the observer. Precise and rapid cultivar identification methods are also required to manage cultivar collections as well as tree peony breeding programs. The objective of this paper is to analyze the discriminatory ability of leaf morphology and Intersimple sequence repeat (ISSR) marker systems for tree peony cultivars. As a result, although there exist large variations of leaf morphology of tree peony cultivars, the morphological characteristics of biternately compound leaves 3, 4, and 5 from the base of a shoot at the middle part of a plant are relatively stable with smaller variations within cultivars (2.7% to 27.1%, 16.8% on average) and with larger differentiations among cultivars (72.9% to 97.3%, 83.2% on average). Statistical and principal components analyses indicate that 12 leaf morphological characteristics are valuable for cultivar classification. ISSR markers present a precisely discriminatory power in tree peony cultivar classification without environmental influences. The cultivars with multiple flower forms, which makes it difficult to make judgment by means of a flower-form-based classification system, have been significantly characterized using leaf morphology or ISSR markers.