Enhancing the efficiency of embryo rescue breeding in seedless grape via direct transformation of abnormal seedlings and secondary embryos

Seedlessness is one of the most valuable agronomic traits in grape. Embryo rescue technology, with its advantages of increasing the proportion of seedless progeny and shortening the breeding cycle, has been widely applied in seedless grape breeding. To improve the efficiency of embryo rescue breeding in seedless grape, this study focused on rescuing and reusing abnormal seedlings generated during the process. By analyzing the influence of parental genotypes from 19 cross combinations on the formation of abnormal seedlings and employing both direct transformation and somatic embryogenesis for their rescue, the study investigated the effects of different abnormal seedling types and parental genotypes on rescue efficiency. Additionally, using the secondary embryos differentiated from the hypocotyls of abnormal seedlings in 3 cross combinations as experimental materials, the aim is to establish a somatic embryo regeneration system. The results revealed that the average proportion of abnormal seedlings (46.25 %) across the 19 cross combinations was significantly higher than that of normal seedlings (33.06 %). Among the abnormal seedling types, leaves without roots and cotyledon albinism were the most prevalent. Using the direct transformation method, the polycotyledonous type exhibited the highest rescue efficiency at 53.06 %, while the cross combination “Perlette × Huozhoucuiyu” achieved the highest abnormal seedling rescue rate of 81.11 %. Furthermore, secondary embryos derived from the hypocotyl of abnormal seedlings were capable of developing into normal seedlings, enabling the rescue of some abnormal seedlings. A cyclic somatic embryogenesis system was established for grape hypocotyls in the crosses “Melissa Seedless × Lüzhoubaoshi” and “Zitian Seedless × Melissa Seedless”. This approach not only significantly enhanced the efficiency of embryo rescue but also expanded the potential applications of abnormal seedlings in somatic regeneration research, providing valuable receptor materials for grape genetic transformation systems.