In the fields of agricultural, food and medical research, the potential impacts of tea on human health are of special interest because tea (Camellia sinensis (L.) Kuntze) is continuously consumed by many people in the world. The evaluation of the diversity of genotypes found in tea germplasm can aid in the improvement of the breeding programme. In this study, the genetic diversity of 30 tea genotypes from two commercial sites of tea production in Iran was investigated by using morphological and 12 ISSR markers. Morphological analysis showed that the diversity between samples of tea was limited, and the narrow matching range was calculated. In cluster analysis at level 0.63, samples were divided into four groups. The application of 12 ISSR primers produced 91 polymorphic bands. PIC test showed a range of 0.41–0.48. Based on the ISSR data, the matching range was obtained in the range of 0.24–0.93. In cluster analysis, samples at level 0.58 were divided into five groups. According to the results, it can be understood that these series of traits and primers can very well recognize genetic differences. Using these markers, genetic diversity was observed among tea genotypes, but this diversity was not such as to be able to separate genotypes of various regions from each other. The results showed that the tea genotypes in Iran had high genetic diversity. As a consequence, the findings of our study will help the development of tea germplasm conservation strategies and their sustainable use in breeding programmes.