The Establishment Of Efficient Detoxification Technology Of Apple Cultivars And Rootstocks

Apple stem grooving virus (ASGV), Apple chlorotic leaf spot virus (ACLSV), Applestem pitting virus (ASPV) and Apple scar skin viroid (ASSVd) were four important appleviruses, which were worldwide distributed in apple production areas, and usually affectedthe growth of tree, and reduce the yield and quality of fruit. There were no effectivemethods of prevention and treatment now, and utilization of virus-free materials was themost effective measure at present. Constant temperature heat treatment was the maintraditional detoxification method. But detoxification efficiency of this method was lower,and varieties of the virus-free seedlings obtained were limited. In order to improve thetraditional detoxification method and raise elimination rate, in this study, different kindsof apple cultivars and rootstocks mainly planted in Yantai area were used todiurnal alternating thermotherapy, chemotherapy and cryotherapy, and the results weredetected by RT-PCR method. Assessed the efficiency of virus elimination by threeprocessing methods, thus the efficient detoxification technology system for differentvarieties of cultivars and rootstocks was constructed. The main results were as follows:1. Through primary culture of different cultivars and rootstocks, to establish theirsterile system, the effect of Hgcl2on the plant was explored. And the optimal processingtime was determined, it was11min for2001, Red General and Yantai Fuji10, and thesurvival rates were more than68%; The sterilization time for Yantai Rootstock1,2and3was12min, and the survival rates were more than70%; For Yantai Fuji8, the time was10min, and its survival rate was90%; For Yantai Fuji6, the sterilization time was13min,and its survival rate was75%.2. The medium formulas suitable for test-tube plantlets in different growth stages hadbeen acquired. Shoot-tip culture medium suitable for2001, Red General, Yantai Fuji8,Yantai Fuji10and Yantai Fuji6was MS+6-BA2.5mg/L+IBA0.5mg/L+5.3g/L Agar+30g/L Sucrose，pH5.8；Shoot-tip culture medium fit for Yantai Rootstock1,2and3wasMS+6-BA2.0mg/L+IBA0.4mg/L+5.3g/L Agar+30g/L Sucrose，pH5.8； Differentiation medium of in vitro plants was MS+6-BA0.5mg/L+IBA0.1mg/L+5.3g/L Agar+30g/L Sucrose，pH5.8；The rooting medium was1/2MS+IAA1.5mg/L+IBA0.15mg/L+5.3g/L Agar+20g/L Sucrose，pH5.8.3. Through the diurnal alternating thermotherapy for Potted seedlings and test-tubeplantlets, there was a find that different cultivars and rootstocks had different hightemperature tolerance. The limit time of2001Potted seedlings and test-tube plantlets wasmore than60d; Red General, Yantai Fuji8, and Yantai Fuji10, their limit time was60d;The limit time of Yantai Fuji6and Yantai Rootstock3was50d; Yantai Rootstock2was40d; The thermostability of Yantai Rootstock1was worst, its limit time was just30d.4. Except that the differentiation rate of Yantai Fuji6and Yantai Roorstock1wasbelow10%, after diurnal alternating thermotherapy, chemotherapy and cryotherapy, thesurvival rate and differentiation rate of shoot-tips of other cultivars and rootstocks werehigh.5. By making a comparison among these three methods, the results showed that thehighest elimination rate of ACLSV after diurnal alternating thermotherapy was83.3%.The rates of ASGV and ASPV after chemotherapy both were100%. Though cryotherapy,the highest elimination rate of ASSVd was75.61%. By combining with different viruselimination efficiencies of different cultivars and rootstocks, the best processingcombinations were accomplished.2001Potted seedlings,diurnal alternating thermotherapy60d (ACLSV)＋chemotherapy60d (ASGV, ASPV)＋cryotherapy (ASSVd); Red GeneralPotted seedlings, diurnal alternating thermotherapy60d (ACLSV)＋chemotherapy60d(ASGV, ASPV)＋cryotherapy (ASSVd); Yantai Fuji8Potted seedlings,diurnal alternating thermotherapy60d (ACLSV, ASPV)＋chemotherapy60d (ASGV)＋cryotherapy (ASSVd); Yantai Fuji10Potted seedlings, diurnal alternating thermotherapy60d (ACLSV)＋chemotherapy60d (ASGV, ASPV)＋cryotherapy (ASSVd);2001test-tube plantlets, diurnal alternating thermotherapy30d (ACLSV)＋chemotherapy60d(ASGV, ASPV)＋cryotherapy (ASSVd); Yantai Roorstock2test-tube plantlets,diurnal alternating thermotherapy30d (ACLSV)＋chemotherapy60d (ASGV, ASPV)＋cryotherapy (ASSVd); Yantai Roorstock3test-tube plantlets, chemotherapy60d (ASGV,ASPV)＋cryotherapy(ASSVd).