Abstract: To effectively identify the varieties of cured tobacco leaves, the genomic DNA of cured leaves of 11 varieties commonly used was extracted by magnetic bead method. The fingerprint spectra based on SSR markers were constructed by PCR and agarose gel electrophoresis. The results showed that five primer pairs with high polymorphism and stability were selected from 24 SSR primer pairs. The amplified products of the five primer pairs were 100-300 bp in length and clear bands were obtained via 3% agarose gel electrophoresis. On the basis of electrophoregram of the five primer pairs, the fingerprint spectra of the 11 tobacco varieties were constructed, and the flow chart of variety identification was drawn for rapid differentiation of the 11 tobacco varieties. This method is simple, practical and convenient to identify the varieties of cured tobacco leaves.