Optimization of efficient virus-free regeneration system from shoot apical meristem of tea chrysanthemum

Abstract [Objective] In order to optimize the efficient virus elimination and regeneration system from shoot apical meristem of tea chrysanthemum, suitable heat treatment duration and detoxification agents, this study aims to improve the problem of low virus-free seedlings obtaining ratio. [Methods] Six tea chrysanthemum varieties were used as materials. The axenic seedlings were pre-treated with alternating heat temperature treatment of 40 ℃/32 ℃ day/night, detoxification agents such as salicylic acid, ribavirin, and abscisic acid, as well as a combination of detoxification agents and heat treatment. The rate of callus induction, regeneration frequency, and average number of regenerated shoots from inoculated shoot apical meristems were compared among different treatment methods, the detoxification efficacy of various treatments were evaluated. [Results] There was a significant difference in the regeneration frequency of shoot apical meristems among different varieties of tea chrysanthemum, with ‘Wuyuan Huangju’ having the highest regeneration frequency of 54.77%. Under different heat treatment days, the difference in regeneration of shoot apical meristems in tea chrysanthemum reached a significant level, with ‘Hangbaiju’ and ‘Qiyuexue’ showing the best regeneration after 40 ℃/32 ℃ heat treatment. Salicylic acid, ribavirin, and abscisic acid treatments inhibited the regeneration of shoot apical meristems. Salicylic acid and ribavirin treatments combined with heat treatment promoted the regeneration of shoot apical meristems, while high concentration abscisic acid combined with heat treatment severely inhibited the regeneration of shoot apical meristems. Treatment with 10 μmol/L salicylic acid combined with heat treatment showed the best regeneration. Virus-free seedlings were obtained through suitable heat treatment combined with detoxification agents. [Conclusion] The combination of 10 μmol/L salicylic acid with 40 ℃/32 ℃ day/night pretreatment for 40 days significantly enhances the regeneration frequency of shoot apical meristems, and obtains virus-free seedlings