Breeding of plants resistant to fungal infections by introduction of a chitinase gene from filamentous fungi.

点击翻译按钮获取中文摘要

Chitinase I of Rhizopus, oligosporus, belonging to zygomycete filamentous fungi, is a pae-pro enzyme. We have cloned chil gene encoding chitinase I.To investigate the effects of expression of fungal chitinase on anti-fungal activity of the transgenic plant, we deleted introns and 3'-region encoding pro-wequence from chil gene and introduced it into some important plants and analyzed phenotypes of the plants. First, the modified chil gene described above was placed between the 35S promoter of Cauliflower mosaic virus and nopaline synthase (Nos) gene terminator and was introduced it into tobacco by the Agrobacterium tumefaciens leaf disc system. Total DNAs were isolated from two of the transformants. Integration of chil gene in these genome DNAs was confirmed by PCR and Southern blot analysis. The expressions of chitinase I in lysates from the young leaves of them were detected by Western blot analysis with anti-chitinase I antibody. Chitinase activities in these lysates were three- to f … More our-fold higher than that from the control plant. Then, the discomycete pathogens Sclerotinia sclerotiorum and Botrytis cirerea were infected on the leaves of them. The symptoms observed with these two transformants were remarkably suppressed as compared with leaves of the control plants. Next, the modified chil gene was placed between the maize ubiquitin gene promoter and Nos gene terminator and was cotransformed into rice with a plasmid containing hygromycin phosphotransferase gene as a selectable marker by the method of particle bombardment. 123 transformants were obtained and regenerated to plants. Total DNAs of them were prepared and used as substrates of PCR analysis and Southern blot analysis. Integration of chil gene was confirmed in 24 transformants. A weak signal of 43 kDa was detected in lysates of the leaves of 4 transformants by Western blot analysis. These transformants were regenerated to plants and infected by Magnaporthe grisea which could cause rice blast disease. The anti-fungal activity of these plants seemed to raise as compared with that of control plants. We also introduced the modified chil gene into tomato and glass and are now trying the regeneration of these transgenic tomato and glass calli. Less