Genetic Stability Assessment of Six Cryopreserved Strawberry (Fragaria × ananassa Duch.) Accessions by Phenotypic and Molecular Studies

SIMPLE SUMMARY: Cryopreservation technology has been developed, demonstrating safe, efficient, and economic benefits for the long-term preservation of genetic resources. It receives more attention because dramatic germplasm losses have been observed in field conservation, caused by many factors such as climate change, natural hazards, and diseases. Therefore, demands for cryobanking crops have been increasing. Cryopreservation methods have been developed for strawberries, the most preferred berry fruit, due to their nutritional and economic values. However, during the cryopreservation process, the explants are exposed to many steps under stressful conditions, which may induce somaclonal variations. The article describes a genetic stability assessment by the greenhouse performance of vegetative and fruit traits and molecular marker analysis for six strawberry accessions. Our results suggest that the cryopreservation protocol used in this study can be applied to implement cryobanking of strawberry germplasm. ABSTRACT: For the long-term preservation of genetic resources, cryopreservation techniques have been developed for strawberry germplasm, mainly using in vitro-grown shoot tips. In this study, genetic stability was tested under greenhouse conditions for six strawberry accessions (IT232511, PHS0132, IT245810, IT245830, IT245852, and IT245860) derived from the following procedures: (1) conventional propagation (GH: greenhouse maintained); (2) in vitro propagation (TC: tissue culture); (3) pretreatment before cryopreservation (−LN: non-liquid nitrogen exposure); and (4) cryopreservation (+LN: liquid nitrogen exposure). To test the performance of phenotypic traits, we measured six vegetative and five fruit traits. There were no distinct differences in most of the characteristics, but a few traits, such as sugar content and pH of fruits in three accessions, showed higher values in +LN compared to GH. However, the differences disappeared in the first runner generation. To test genetic variations, a total of 102 bands were generated by twelve inter simple sequence repeat (ISSR) primers. A few polymorphic bands were found only in plants derived from TC of IT245860, which was not cryopreserved. The sequencing analysis of four polymorphic bands produced by ISSR_15 showed that none of these sequences matched the characterized genes in NCBI. Phenotypic abnormality was not observed across all plants. This study indicates that cryopreserved plants of the six strawberry accessions are phenotypically and genetically stable. Therefore, the results of this study can help to implement cryobanking of strawberry germplasm.