Abstract

Potato (Solanum tuberosum L.) is one of the most important crops grown in Norway, and virus-free plants are required for commercial potato production and for preservation of potato germplasm. The present study evaluates three in vitro therapies – meristem culture, cryotherapy, and chemotherapy combined with thermotherapy – to eliminate viruses from eight historically valuable potato cultivars belonging to the Norwegian potato germplasm. Potato virus Y, potato virus M, potato virus X and potato virus S were present in eight selected old potato cultivars due to long-term conservation in open field. Double-antibody sandwich enzyme-linked immunological assay (DAS-ELISA) and biological indicators were the standard tests used to confirm virus infection in our study. Six virus-free plants from four potato cultivars were obtained after meristem culture, and no virus-free potato cultivars were obtained after cryotherapy. Virus-free frequency for eight different potato cultivars after combining chemotherapy with thermotherapy varied from 36.4% to 100%, with single virus elimination rates of between 74.2% and 92.9%. Chemotherapy compared with thermotherapy was the most effective of the three in vitro therapies used in this study. Highly sensitive small RNA high-throughput sequencing (HTS) was used to evaluate the virus status of potato virus-free materials after virus eradication, and no virus was found, which was consistent with the results of DAS-ELISA and biological indicators. Small RNA HTS has been reported for the first time to evaluate the virus status after virus elimination and to control virus-free potato nuclear stocks.

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Abstract

Virus diseases have been a great threat to production of economically important crops. In practice, the use of virus-free planting material is an effective strategy to control viral diseases. Cryotherapy, developed based on cryopreservation, is a novel plant biotechnology tool for virus eradication. Comparing to the traditional meristem culture for virus elimination, cryotherapy resulted in high efficiency of pathogen eradication. In general, cryotherapy includes seven major steps: (1) introduction of infected plant materials into in vitro cultures, (2) shoot tip excision, (3) tolerance induction of explants to dehydration and subsequent freezing in liquid nitrogen (LN), (4) a short-time treatment of explants in LN, (5) warming and post-culture for regeneration, (6) re-establishment of regenerated plants in greenhouse conditions, and (7) virus indexing.