Switchgrass (Panicum virgatum L. ) is considered to be one of the ideal bioenergy plants. The genetic variation and relationship of 96 switchgrass germplasms were analyzed by SCoT (start codon targeted polymorphism) molecular markers. 20 SCoT primer pairs were used to amplify SCoT fragments for 96 switchgrass germplasms. The primers generated a total of 445 bands among which 402(90.31%) bands were found to be polymorphic. The value of locus polymorphism information contents (PIC) ranged from 0.166 to 0.410, with a average of 0.332. Marker index is 10.20. The genetic similarity coefficient among the germplasms was ranged from 0.498 to 0.912, and the average content was 0.688, indicating that the genetic variation could be revealed using SCoT markers. 96 switchgrass germplasms were clustered into two groups (Upland and Lowland) by the cluster analysis. The results of POPGENE version 1.32 analysis indicated that the genetic diversity index(H) of 96 was 0.406, and the index of Shannon(I) was 0.271, indicating that there was comparatively great genetic diversity and high genetic variation among the switchgrass germplasms. Analysis of molecular variance(AMOVA 1.55) indicated that contribution(72.85%) of variability came within population level and account for 27.15% of total variability. The results indicated that SCoT markers were feasible and effective to analyze the genetic diversity of switchgrass and the results will facilitate the germplasms of switchgrass exploitation and utilization.