Iran has many valuable genetic resources of medicinal plants. It is important to exploit a variety of genes from this variation. Considering the importance and various uses of medicinal plants, today many aspects of plant science researches have focused on the use of these plants. German chamomile ( Matricaria chamomilla ) is an herbaceous plant, perennial, native to the Mediterranean region and Asteraceae family. It is native to Mediterranean regions and some researchers reported Asia as its origin. This study evaluated the genetic diversity of 21 Iranian and two European populations using ISSR molecular markers and morphological characteristics. According to the results, out of 20 ISSR primers 10 one generated polymorphic bands with clear banding patterns. A total of 208 bands were generated in which 193 were polymorphic bands (92.78% of the variation). Bands were scored as zero for absence and one for presence. The genetic similarity coefficient was calculated based on Simple Matching (SM) (0.92). Results of cluster analysis and principal coordinate analysis (PCoA) of the chamomiles were divided samples into four groups. Analysis of molecular variance (AMOVA) showed more variation within groups (58.98%) compared with the variation among groups (41.02%). Based on the collection sites three groups were assumed for AMOVA analysis including Centre (C), North- West (NW) and South- East (SE). Most of the genetic diversity was obtained for Center (0.25) and the lowest one was observed in South East (0.054), respectively. Cluster analysis based on morphological characters divided genotypes into two major groups. Totally, there were no correlation between morphological and molecular results. Finally, Acceptable variation was observed in molecular and morphological results for chamomiles and it is beneficial for future breeding programs. Keywords: Matricaria chamomilla , Molecular marker, Morphology