Elimination of Potato Viruses (PLRV and PVY) by Cryopreservation of In Vitro Grown Shoot Tips of Potato (Solatium tuberosum L.)

摘要

Cryopreservation has been recognized as a practical and efficient tool for long-term storage of vegetatively propagated plants. This study was conducted to investigate the effect of sucrose concentration and hardening temperature on the survival of cryopreserved shoot tips and virus elimination through cryo-therapy. Virus-free in vitro plantlets of potato cultivars 'Atlantic' and 'Superior' were used for survival studies whereas naturally infected in vitro plantlets of potato genotype T9-99' by potato leaf roll virus (PLRV) and potato virus Y (PVY) were used for virus elimination by Cryopreservation. Excised shoot tips were cryopreserved by vitrification using plant vitrification solution-2 (PVS-2). Nine-percentage sucrose concentration gave the highest survival of 41.7% in 'Atlantic' and 33.3% in 'Superior'. The most optimum hardening temperature for the highest survival of 'Atlantic' (43.3%) and 'Superior' (32.3%) was 10°C. The virus status of the regenerated in vitro plantlets before and aftertreatment was tested by employing double antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) and reverse transcription polymerase chain reaction (RT-PCR). Virus status was subsequently detected following greenhouse acclimatization by both methods. In the case of virus elimination using different steps of Cryopreservation, none of the steps were found effective before freezing. The conventional meristem tip culture resulted in 13.6% PLRV and 11.4% PVY free plantlets, while the cryoprotectivetreatment resulted in 36.8% PLRV free plantlets and 42.1% PVY free plantlets. Cryopreservation steps were not affected for virus elimination, whereas in the case of Cryopreservation procedures, vitrification and Cryopreservation weife' more effective than the encapsulation-vitrification and Cryopreservation for elimination of PLRV and PVY from the in vitro plantlets. Leaf and tuber morphology of potato plants regenerated after Cryopreservation (cryo-therapy) was similar to that of the control. Thus, this study demonstrated that PLRV and PVY can be successfully eliminated from the infected in vitro shoot tips of potato by cryo-therapy.