Research On Cry3Aa Transformed Into Populus×Euramericana’Guariento’ And Populus×Euramericana’2001’

It is practical to improve resistant ability for current poplar cultivar species toLonghorn beetles. Populus×euramericana ’Guariento’ and Populus×euramericana’2001’ are good candidates to improve the resistant ability for their fast-growing andhigh-yield. Optimized plant regeneration system was studied in this dissertation, andCry3Aa was tried to transform into Populus×euramericana ’Guariento’ and Populus×euramericana ’2001’.The preliminary conclusions from the research are as followings:1. The optimum pH of the culture medium is6.3. In this test, the pH of mediumreduced0.5after sterilizing. The best basic media pH for the two explants Populus×euramericana ’Guariento’ and Populus×euramericana ’2001’ was6.3. The mediumpH adjusted to6.8was the most appropriate before sterilization.2. Populus×euramericana ’Guariento’ and Populus×euramericana ’2001’callus induction pHenotype consistency. The most suitable medium: MS+2,4-D2mg/L. After comparing the size, color, texture and increment of callus, MS+2,4-D2mg/L induced the largest callus quantity at the same raise time. The basal mediumwas MS medium, added with2,4-D2mg/L, the callus rate of Populus×euramericana ’Guariento’ was60.07%, Populus×euramericana ’2001’ was40.65%;Added with6-BA1mg/L and2,4-D2mg/L, the callus rate of Populus×euramericana ’Guariento’ and Populus×euramericana ’2001’ were36.67％,29.67％respectively. Therefore, MS+2,4-D2mg/L was selected as the callus proliferationmedium in this study.3. Callus induction medium for optimum bud: MS+KT0.1mg/L+6-BA1mg/L. Itcould tighten the Populus×euramericana ’Guariento’ and Populus×euramericana’2001’ loose callus gradually, and then the small buds. Unconsolidated callus induction budding is not easy, the appropriate hormone combination with the appropriatehormone concentrations in the suitable artificial environment is important. Thegermination rate of Populus×euramericana ’Guariento’ and Populus×euramericana’2001’ on the MS+KT0.1mg/L+6-BA1mg/L medium were53.3%,48.65%, on the48+6-BA1mg/L+KT1mg/L medium were17.95％,17.86％, and on the MS+NAA0.05mg/L+6-BA1mg/L+KT0.1mg/L medium were4.76%,5.88%respectively.4. The critical kanamycin concentration to restrain explants differentiation wasdetermined. As the poplar stem material, the concentration gradient of kanamycin was30mg/L,50mg/L,60mg/L,70mg/L,80mg/L,90mg/L,100mg/L, with noadded stems inoculated media sheet as CK, each gradient inoculated30stem thinsections, after25d training and observation, the callus rate of Populus×euramericana’Guariento’ and Populus×euramericana ’2001’ on the medium with kanamycin50mg/L were10.53%,18.18%; and on the medium with kanamycin60mg/L were5.00%,9.38%; all materials bleaching and dying on the medium with kanamycin70/Lor more. Callus screening process and gradient settings with same stems. Therefore,kanamycin70mg/L is the appropriate concentration for the research of stem segmentsand callus screening. While study poplar petiole,leaf material,kanamycin concentration50mg/L is suitable for screening concentration.After testing conditions, to geneticallymodified materials, callus highest conversion rate reached80.00%, the lowest38.18%for Populus×euramericana ’Guariento’, and the highest reached93.75%, the lowest21.68%for Populus×euramericana ’2001’. But to Populus×euramericana’Guariento’ and Populus×euramericana ’2001’, the direct organogenesis conversionefficiency is very low, not exceed0.5‰. Therefore, through transformation of callusway transgenic can greatly improve the efficiency of genetic transformation forPopulus×euramericana ’Guariento’ and Populus×eramericana ’2001’.