Essential molecules are embedded within the millenary crop Tropaeolum tuberosum (mashua); these compounds are critical for the Andean people’s traditional diet and extensively utilized by the pharmaceutical industry in Peru. In the Andean region, conventional cropping techniques generate microtubers susceptible to a viral infection, which substantially endangers mashua’s production. Therefore, we developed an innovative in vitro technique condition for enhancing the agriculture process for micro tubers production. The temporary immersion system (TIS) permits the production of high-quality microtubers in a reduced space, a lower amount of time, and in large quantities compared with tubers grown under traditional conditions. To obtain T. tuberosum’s microtubers via TIS, we propagated seedlings, utilizing TIS-RITA® vessels. A set of immersion frequency times were evaluated. Interestingly, results showed that immersion at 2 min every 3 h was more beneficial compared with 2 min every 5 h based on microtubers produced after 10 weeks from the treatments, revealing an efficient frequency setting which outputted improved microtubers quality and production.