Inhibition Of Arabidopsis AZI1 To The Growth Of Saccharomyces Cerevisiae And Infection Of Botrytis Cinerea Pers.ex Of Garlic Sprout

Resistance of Arabidopsis AZI1 to fungus was analyzed with Saccharomyces cerevisiae expression vector pESC and plant expression vector pPZP211. Compared to yeast cells transformed with empty vector, the growth ability of yeast cells harbouring pESC-AZI1 declined obviously on SC-URA medium containing 2% sucrose. On SC-URA medium supplemented with 2% galactose, yeast cells harbouring pESC-AZI1 could not form colony after 48h incubation at 28℃, while only a few colonies could be found after 144h. Western blot analysis showed that yeast cells harbouring pESC-AZIl could express AZI1 efficiently after induction with galactose. The spores of Botrytis cinerea Pers.ex of garlic sprout were used to infect the leaves of wild-type Col-0 and AZI1 overexpressing transgenic plants. Generation of H2O2 was analyzed by DAB staining 24h after inoculation. In Col-0 wild-type plants, the staining colour of infection sites on leaves were a little lighter, it showed that only small amounts of H2O2 were produced at the infected sites of leaves and the pathogens could spread to surrounding cells. In contrast to this, the infection sites on leaves of AZI1 overexpressing plants were stained intensively by DAB, which would accumulate large amounts of H2O2, indicating the transformants could prevent the spread of pathogen by the death of local cells in infected sites. Trypan blue staining showed that a large number of cells in pathogen infected leaves of wild-type Col-0 plants were died in comparison with the leaves of AZI1 overexpressing plants. Besides, the expression of AZI1 could be induced by salicylic acid and peaked after 24h in Col-0 wild-type plants. All these results suggested that AZI1 play an important role in defense system of Arabidopsis against biotic stresses. In addition, the following three aspects of researches were carried out in this work.1, prokaryotic expression vectors pET32a-AZI1 and pET32a-EARLI1 were constructed and transferred to BL21(DE3) strain of Escherichia coli successfully.2, Plant fusion expression vector pCAMBIA1302-AZI-GFP was constructed and introduced into LBA4404 strain of Agrobacterium tumefaciens which was used in genetic transformation of the wild-type Arabidopsis thaliana ecotype Col-0 and Nicotiana tobacum genotype QinYan95 subsequently. 3, Agrobacterium tumefaciens strain ABI containing pPZP211-AZI1 was adopted in transformation of Nicotiana tobacum genotype QinYan95,11 putative transgenic plants were preliminarily verified by PCR and RT-PCR.