A simple mid-term preservation method (SMPM) of plant callus under low temperature conditions

Abstract

The repeated monthly or weekly subculture of plant callus is labor intensive and increases the risk of somaclonal variation from the parental callus line. The most effective method for preserving plant callus is cryopreservation, which involves storage in liquid nitrogen. However, this method cannot be applied to the callus of different plant species in the same manner, so it is difficult to develop a standardized cryopreservation method. In addition, the survival rate of the frozen callus after thawing and the regeneration rate after survival are uncertain. Therefore, it is necessary to develop a method to extend the subculture interval of plant callus in an active state. In this study, active plant calli of various species without freezing was incubated at 15°C for 4 to 12 weeks without subculture. After 12 weeks, 8 lines of plant callus grew less than 2-fold when cultured at 25°C, but at least 2 times as much when cultured at 15°C. Moreover, total antioxidant activity did not differ significantly between plant callus recovered at 25°C after culturing at 15°C or at 25°C. These results show that the subculture interval can be extended at a temperature of 15°C without need for modified medium composition or additional processes. In addition, positive results in all calli of several plant species are expected to reduce labor as well as somaclonal variation by increasing the subculture.