”Cat ginseng”, the dried root of Actinidia macrosperma, is a famous traditional Chinese medicine against cancers in eastern China. The roots of some other species of the genus Actinidia such as A. valvata and A. melanandra have also been used as fake ”cat ginseng”, but they have little therapeutic value. However, identification of the original plants used to make the crude drugs is difficult, especially during the vegetation period. In this study we developed molecular markers for the determination and authentication of A. macrosperma. The restriction digestion of the chloroplast trnK region using endonucleases DdeI and DraI produces two unique patterns in A. macrosperma, and in the matK sequences, there are 11 sites unique to A. macrosperma. The molecular markers provide an effective and accurate identification and authentication of A. macrosperma in Actinidia.