Establishment of an Efficient Plant Regeneration System for the Elite Clone DH32-28 of Eucalyptus urophylla×E.grandis
This study investigated the effects of plant growth regulators on the elongation of internodes in proliferated seedlings and plant regeneration using aseptic seedlings from three elite clones of Eucalyptus urophylla×E.grandis(DH32-26,DH32-28 and DH32-29).The effects of clones,plant types and explant types on plant regeneration capability were further studied.The easily regenerated clone DH32-28 was used as the research material to further investigate the effects of plant growth regulators and callus induction time on shoot regeneration.The results showed that the clone was the primary factor influencing the in vitro regeneration in E.urophylla×E.grandis.The regeneration efficiency of DH32-28 was significantly higher than those of DH32-26 and DH32-29.The internodes of all proliferated seedlings exhibited significant elongation after treatment with GA3 or IBA,and the GA3-treated DH32-28 seedlings had the highest shoot regeneration rate and the highest number of regenerated shoots.Significant differences were observed in the shoot regeneration rates among different types of explants,with the highest regeneration rate found in the stem segments of elongated seedlings of DH32-28,followed by petioles and leaves.The optimal conditions for DH32-28 in vitro shoot regeneration were as follows.The internode stem segments of elon-gated seedlings after 20 days of GA3 treatment were used as explants.The callus-inducing medium is 1/2 MS supplemented with 0.22 mg/L TDZ,0.019 mg/L NAA,30 g/L sucrose,and 7 g/L agar with a 20 days incubation.The optimal shoot-inducing medium is 1/2 MS supplemented with 0.25 mg/L 6-BA,0.125 mg/L NAA,30 g/L sucrose,and 7 g/L agar,and the shoot regeneration rate reaching 95.00%after 30 days of induction.The regenerated shoots were placed on root-inducing medium 1/2 MS supplemented with 0.5 mg/L IBA,30 g/L sucrose,and 7 g/L agar for root induction,where all regenerated shoots established roots.This study successfully established an effi-cient plant regeneration system from stem segments of the excellent clone DH32-28 of E.urophylla×E.grandis,laying the groundwork for the establishment of genetic transformation systems and subsequent investigations into gene function and genetic engineering.
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