Effect Of Thermotherapy And Chemotherapy On Viruses In In Vitro Pear Plants

Apple stem grooving virus (ASGV) and Apple chlorotic leaf spot virus (ACLSV) are two important viruses in pear, which are worldwide distributed. In commercially cultivated pear, these two viruses usually do not cause visible symptoms, but they decrease the growth and productivity of infected trees. One of most effective measures for the controlling of these virus diseases is utilizing certified healthy propagation materials. Thermotherapy and chemotherapy are the most widely used measures for the production of virus-free plant materials. In these studies, for obtaining higher virus elimination efficiency, the virus distribution features in in vitro pear plants during thermotherapy and chemotherapy were analyzed in this study. Seeking results are listed as following.1. 20 in vitro plant clones of Pyrus pyrifolia cv.'Huanghua'were detected by ELISA and RT-PCR for ASGV and ACLSV. Results showed that 13 clones were infected by both ASGV and ACLSV. These clones were propagated on MS medium for the following studies.2. Tissue printing immunoassy was used for the detection of ASGV and ACLSV in different parts of stems of in vitro pear plants. Non-specific reaction was decreased by improving the absorbance protocols of polyclonal antibodies against these two viruses, prolonging the periods of washing steps. The distribution of viruses in vertical sections of plants could be easily observed based on the present of purples products. Results showed that both ASGV and ACLSV had a high concentration in the base and tip of these plants, and the intermediate section also had virus distribution with a lower amount.3. Biotinylated cDNA probes specific to ASGV and ACLSV were prepared from cloned fragments. The dot-blot hybridization and tissue-printing hybridization methods were studies for the detection of these viruses in plant crude extracts and tissues, respectively. These viruses in both vertical and horizontal sections of the stem of in vitro plants were detected by in situ tissue-printing hybridization. Similar results were achieved as tissue printing immunoassy, but with a more intensive reaction-signal.4. In vitro plant clones of Pyrus pyrifolia cv. 'Huanghua' were treated under 37℃ and 32℃/39℃ (8h /16h alteration) respectively. Changes of virus titers in stems of treated plants during thermotherapy were analyzed by tissue printing immunoassy and in situ tissue-printing hybridization established formerly. Results indicated that virus titers showed a decreasing tendency while the treatment periods were prolonged, and the tendency was more obvious in shoot tips than bases. When plants were treated for 35 days under 37℃ or 50 days under 32℃/39℃, no purple signal was observed in shoot tips,...