Identification Of Resistance Of Tea Use Chrysanthemum To Fusarium Wilt And Mechanisms Underlining The Resistance

Chrysanthemum(Chrysanthemum morifolium Ramat.)has a history of more than 1600 years of application.Chrysanthemum is one of the top ten famous flowers in China and one of the important flowers in garden applications.Chrysanthemum for tea can be used to moisturize the throat,and has the functions of clearing away heat,eyesight and hangover.Chrysanthemum for tea wilt is a soil-borne disease caused by Fusarium oxysporum.In recent years,the damage of chrysanthemum for tea wilt is serious.In the perennial continuous production area of Chrysanthemum for tea,the susceptibility rate of chrysanthemum increased,the yield decreased and the quality decreased.It has become an important issue to be solved in the production of chrysanthemum for tea.The only way to solve this problem is to screening excellent germplasm,clarifying the mechanism of disease resistance,and cultivating excellent varieties of chrysanthemum for tea.However,there are few studies on the resistance of Chrysanthemum Fusarium wilt.In this study,the pathogenic bacteria were isolated from the diseased plants which collected from the’Fubaiju’ production area,and the pathogenic strain was identified as Fusarium oxysporum by morphological observation and molecular identification.The 33 germplasm resources preserved in the "Chinese Chrysanthemum Germplasm Resources Conservation Center"were artificially inoculated and treated by seedlings to study the distribution of resistance to F.oxysporum,and the anti-inductive materials were screened;Study the physiological and biochemical differences of the materials to explain the disease resistance mechanism;Through protoplast transformation,the plasmid containing the gfp fragment was transferred to M15 to obtain a transformant inoculation material containing the gfp fragment,observing the dynamic process of infecting plant roots and laying the foundation for the study of molecular mechanisms such as transcriptome sequencing.1.The diseased plant is from ’Fubaiju’ production field in Futianhe Town,Macheng City,Hubei Province.The pathogenic bacteria were isolated by tissue block separation and the monospora strain was cultured to purification.The isolated strains were identified by morphological and molecular biological identification methods combined with the construction of phylogenetic trees.The pathogenicity of the obtained strain was tested,and the pathogenic strain was identified as Fusarium oxysporum.By studying the effects of different concentrations of spore suspension on the incidence of ’Fubaiju’,it was found that the spore concentration was 1×106！mL-1,and the inoculation effect Cas the best.2.Under greenhouse conditions,identification of F.oxysporum resistance in 33 chrysanthemum for tea varieties by artificial inoculation of strain M15 at seedling stage.Observe the statistical disease tree and the disease tree,calculate the incidence rate and disease index,and carry out resistance classification.Screened for two high-resistance variety ’Qiyuebai’(HR,DSI:0～15),nine resistant varieties such as‘Hangbaiju’(R,DSI:15～30),’Suju 7 hao’,seventeen medium-resistant varieties such as ’Chuju’ and ’Boju’(MR,DSI:30～50),four susceptible varieties such as ’Fubaiju’ and ’Huangshangongju’(S,DSI:50～70),and one high-sensitivity variety‘Huangju’(HS,DSI:70～100).3.The leaves and roots of the resistant variety ’Suju 12’ and the susceptible variety’Huangshan Gongju’ were selected at 0 d,1 d,2 d,4 d,6 d,8 d,12 d,24 d,36 d respectively.The determination of the defensive enzyme activity of the sample.And the experimental results showed that PAL activity,PPO activity,CAT activity and POD were negative correlated with the incidence of chrysanthemum for tea,SOD activity may not have a significant relationship with the occurrence of blight.4.The plasmid carrying the gfp fragment was transformed into M15 oxysporum)by protoplast transformation.Successfully obtained a mutant strain of F.oxysporum that stably and continuously expressed green fluorescent protein,Identification of resistance to transformants indicated that constitutive expression of gfp did not affect the pathogenicity of M15.The process of attachment,infection and colonization of pathogens in tea chrysanthemum was observed by an upright fluorescence microscope.Spores on the root surface begin to germinate on the day of inoculation and the hyphae adhere to the longitudinal axis of the root surface.