Regeneration Of Three Genotypes Of Malus Germplasm Resources

Malus toringoides(Rehd.)Hughes has the characteristics of apomixis,and it is an excellent resource of apple rootstock.Malus ’Winter Red’ and Malus ’Radiant’ are species with high ornamental value and widely used as ornamental plants in North America.They contain special dihydrochalcone compounds.In vitro regeneration of apple is progressing rapidly and has been successfully cultivated for many varieties and rootstocks.However,for apomictic apple rootstocks,only Malus hupehensis var.pingyiensis and Malus xiaojinensis have established complete regeneration and genetic transformation system.In addition,embryo rescue has made good progress in seedless grape cultivation,but there is little research on embryo rescue in apple.In this study,Malus toringoides(Rehd.)Hughes,Malus’ Winter Red’ and the hybrid progenies of Malus ’Winter Red’ and Malus ’Radiant’ were used as experimental materials to establish a regenerative system through tissue culture technology and embryo rescue,in order to lay a foundation for later genetic transformation.The main experimental results are as follows:1.Through flow cytometry determination,it was found that M.toringoides(Rehd.)Hughes was triploid,while M.’Winter Red’ and M.’Radiant’ were diploid.2.It was proved that the pollution rate of the established explants on the day of sampling was lower than that on the day after sampling.In addition,when disinfecting,it is better to wash with 75% ethanol for 1.5 min,then wash with sterile water once,then wash with 8% Na Cl O solution for 8 min,wash with Tween for 12 min,and finally wash with sterile water for 3 times.3.In the process of subculture,when the concentration of sodium thiosulfate in the medium was 0.5 mg/L,the concentration of IAA was 0.2 mg/L,and the concentration of6-BA was 1.0 mg/L,1.5 mg/L,2.0 mg/L and 2.5 mg/L,the plants grew well.Adding0.1mmol and 0.5mmol sodium ascorbate solution to leaf disc culture medium helped to reduce leaf Browning.4.Through the establishment of regenerative system,the optimal medium for subculture of Malus ’Winter Red’ was 4.43 g/L MS+30 g/L sucrose +7.5 g/L AGAR +0.3 mg/L6-BA+0.9 mg/L IAA.The stem height was significantly higher when the plants were placed horizontally at an Angle of less than 30 degrees than when the plants were placed vertically.The optimal medium for foliar disc proliferation was 4.43 g/L MS+30 g/L sucrose +7.5 g/L AGAR +1.0 mg/L TDZ+0.3 mg/L NAA.The suitable rooting medium was 4.43 g/L MS+20g/L sucrose +7.5 g/L AGAR +0.5 mg/L IAA+0.5 mg/L IBA.5.Through embryo rescue of the hybrid progenies of Malus ’Winter Red’ and Malus’ Radiant’,it was concluded that the hybrid progenies were suitable for embryo rescue 60 days after pollination.The optimal subculture medium was 4.43 g/L MS+30 g/L sucrose +7.5 g/L AGAR +0.3 mg/L 6-BA+0.9 mg/L IAA.The optimal medium for foliar disc proliferation was 4.43 g/L MS+30 g/L sucrose +2.5 g/L phytogel +7.5 g/L AGAR +5 mg/L 6-BA+1 mg/L TDZ+0.3 mg/L NAA.The suitable rooting medium was 4.43 g/L MS+20 g/L sucrose +7.5g/L AGAR +0.5 mg/L IAA+0.5 mg/L IBA.