Abstract: Optimization of DNA isolation protocol for genetic characterization of any medicinal plant is the necessary and primary step. Plant leaves of medicinal plants are rich in secondary metabolites, polysaccharides and polyphenols that are problematic during isolation of genomic DNA. Besides, succulent plants freezed under liquid nitrogen turn to stonelike tissues that are difficult to grind. Flowers were not considered as a source of genomic DNA as the case of young leaves, this might be due to the seasonal nature and short half-life feature. Several protocols described for plant DNA isolation from Aloe barbadensis Miller using leaf, found difficult and tedious. This plant contains exceptionally high amount of secondary metabolites that interfere with DNA isolation.