Studies On Tissue Culture And Genetic Transformation Of Sand Pear

Pear (Pyrus spp.) is one of the most important fruits in the world and also a famous fruit in China. Excellent quality, attractive color, good taste and marvelous fragrance make it a poplular fruit in the market. Sand pear (Pyrus pyrifolia Nakai) cultivars also have other characteristics such as earlier fruit ripening, anti-heat and anti-fire blight. But most cultivars of sand pear still have a lot of shortages such as sensitive to frost and diseases, bad apperance of fruit and so on.With the development of biotechlolgy, plant gene transfer technique has been used far and widely. So It appears promising that breeding fruit new cultivar with excellent traits by biotechlolgy. So this research described sand pear tissue culture and gene transformation technique. Shoots were regenerated from the young leaves of sand pear cv. 'cuiguan', 'xueqing'. Based on regenration system, the condition of Agrobacterium-mediated transformation were explored. The results were summarized below.1. Adventitious shoot regeneration from in-vitro leaves of sand pear cv. 'Cuiguan', 'Xueqing' and ' Xizilv' was studied. Among the three cultivars, 'Cuiguan' showed the relatively higher response to regeneration, 'Xueqing' relatively lower, and no response for ' Xizilv' . Dark treatment was necessary for regeneration of 'Cuiguan' leaves in the experiment. When NAA concentration was fixed and BA concentration was changed from 3 mg/L to 7 mg/L, Adventitious shoot regeneration rate from in-vitro leaves of 'Cuiguan' was increased steadily. The effect of TDZ on regenration from in-vitro leaves of 'Cuiguan' was very small. MS was optimal medium on regenration from in-vitro leaves of 'Cuiguan'. AgNO3 enhanced the regeneration efficiency of leaves while its concentrations varied from 0.1 mg/L to 0.5 mg/L.2. Scanning electron microscopy (SEM) observation on 'Cuiguan' revealed that meristem-like structures were already visible at day 10 after the initiation of the experiment and well-developed shoots were observed after two months. SEM also showed that more meristems were observed than the shoots eventually formed from them.3.Through experiments of the effect of Cef on callus induction and shoot regeneration from leaves of 'Cuiguan' and the effect of Kan on callus induction, optimal concentrations of Cef and Kan were established, 200 mg/L and 25 mg/L respectively. About 1000 pieces of leaves of 'cuiguan' were infected by Agrobacterium trmefaciens, but resistant shoots have not been obtained yet for low regeneration rate. The effect of mannose on callus induction and shoot regeneration from leaves of sand pear was investigated. The result implied that mannose cannot be uitilized by these three cultivars. So precondition of PMI resistant system was possessed.For the first time this research described time-course changes of regeneration from leaves of sand pear examined under scanning electron microscope. It can help us to understand dynamic course of ogan development morphologic in detail and grope for best ways to increase regeneration frequency.Progress of genetic transformation of pear tree is detained by great difficulties, papers on pear transformation still have not been published in homeland, while abroad a few papers related with pear transformation were only limitted on cultivar of Europe pear and pea pear. In this study, the conditions of genetic transformation of sand pear were investigated first and optimal concentration of antibiotic on leaves of 'cuiguan' was established. This may be referred by other cultivars of pear.