Abstract: The genetic similarity coefficients and cluster analysis maps on Lactuca pulchella (Lp) and its 11 closely related species were obtained by using PCR to amplify ITS, matK and rbcL gene from the total DNA fragments of the lettuce varieties. Based on the ITS sequences, a similarity between Lp and L. sativa var. ramose was found to be 93.6%; between Lp and L. sativa , 93.1%; between Lp and L. serriola , 92.6%;and, between Lp and Mulgedium tataricum , the lowest at 87.8%. According to the matK sequences, a similarity between Lp and L. sativa var. ramose was shown to be 99.2%;between Lp and L. sativa , 99.0%, ; between Lp and L. serriol , 99.1%; and, between Lp and M.tataricum , 98.3%. It appeared that Lp was related closer to the Lactuca species than Mulgedium . By comparing the rbcL , Lp and L. sativa L. var. ramosa had a perfect match with a 100% similarity; whereas, Lp and L. sativa , L. indica , L.serriola , L. hirsute or L. perennis , at 99.7%;and, Lp and M. tataricum , at 99.4%. Consequently, it was concluded that Lp belonged to the genus Lactuca .