The microtuber is considered one of the most effective means of spreading basic materials, as well as transporting and preserving potato germplasm varieties. To define the optimal conditions for the potato microtuber in vitro germination of Aida, Atlas and Odessa varieties, the effects of temperature, physiological age and grade (size) were evaluated. The study conducted at three different temperature levels has demonstrated that the most favorable temperature for microtuber germination at a higher and faster germination rate was 25℃, regardless of the variety. In addition, microtubers of large caliber, greater than 4 mm, germinate more quickly, with a higher germination rate, than smaller size ones (less than 4 mm) for all genotypes. For Atlas, Aida and Odessa varieties, a germination rate equal to 86.66%, 70% and 70% respectively, was obtained for microtubers with a caliber superior to 4 mm. Physiological age influences microtuber germination. The mean length of sprouts, reached after a 7 week incubation period, was more marked at “multiple sprout” and “branched sprout” stages than at a “monosprout” stage. The average length was 2.35 cm, 2.48 cm and 1.5 cm, respectively. Thus, it is necessary to plant microtubers at a “multiple sprout” stage to optimize their yield in plants and minitubers.