Johnsongrass [Sorghum halepense (L.) Pers.] is a malignant weed in the world, threatening biodiversity at invaded habitats in more than 50 countries. Because of similarity in morphological characters, S. halepense and its relatives, S. almum, S. nitidum, S. propinquum, S. sudanense, and S. bicolor, etc. was difficult to identify. As a supplementary methodolgy for morphology identification, a molecular detection method was established. Sequence Characterized Amplified Regions (SCAR) marker is a recent established, reliabile, and stable molecular marker based on RAPD maker, an effective way for germplasm identification. In this study, one specific band of S.halepense was screened by 163 pairs of RAPD primers. According to the sequences of the band, a pair of special SCAR primers SH1/SH2 was designed and verified by 65 Sorghum DNA samples from all over the world. The results showed SCAR primers SH1/SH2 can be used to distinguish S.halepense and its relatives rapidly with three exceptions of Australia geotypes.