Identification Of Pathogens From Lotus,Control Technigues And The Resistance Gene Screening Of Lotus Rot Disease

Lotus,an economically important aquatic plant,is widely planted in China due to its high edible value and medicinal value.In the process of lotus planting,its yield is affected by many factors.The white lotus lot disease has emerged as a major threat to white lotus in Jiangxi province,and it has become a bottleneck problem that restricts the cultivation of white lotus.In this paper,pathogens of white lotus diseases were investigated and identified,the fungicide screening of the pathogens were acrried out.The field experiments were conducted to control the white lotus rot disease.In addition,RNA-seq was performed on the infected lotus stem（LD）with Fusarium oxysporum and healthy lotus stem（LH）samples,the transcriptome differences between the infected plant and the healthy plant were systematically analyzed.Two significant differentially expressed agmatine-coumaryl acyltransferase-related genes were obtained.And functional analysis of two genes were conducted.It was used to excavate the white lotus rot disease resistance gene in molecular level.The results were as follows:1.From 2017 to 2019,a survey of white lotus diseases was conducted in Jiangxi Province.8 lotus leaf diseases were identified through morphological indentification,pathogenicity,and molecular biological,and they are Nigrospora sphaerica,Bipolaris sp.,Corynespora cassiicola,Lasiodiplodia theobromae,Curvularia trifolii,Fusarium equiseti,Neofusicoccum parvum,Phomopsis sp..2.The toxicities of 5 fungicides against 7 kinds of white lotus leaf disease pathogens were tested by the mycelial growth rate method.The results showed that the 7 pathgens were well supressed by prochloraz,pyraclostrobin,benzpropiconazole,and the biological agent Bacillus subtilis.The EC₅₀ values of these fungicides were all lower than 10 mg/L.Different combinations of soybean,soybean ferment,carbendazim,formosa,and microbial fertilizers were used to perform white lotus rot disease prevention and control.The results showed that incidence of the used soybean and soybeanferment field was reduced by 50%-70%,the yield of lotus seeds was 2-3 times higher than control,and the weight of a single lotus seed was increased by 15%compared with control.3.The transcriptome analysis on the infected lotus stem tissue（LD）of Fusarium oxysporum and uninfected healthy lotus（LH）was conducted.When FPKM>1,47.96%of the genes in the LD group had varying expression changes.Significant differences were observed in the expression levels of genes related to phenylpropane biosynthesis,α-linolenic acid metabolism,glutathione metabolism,phenylalanine metabolism,flavonoid biosynthesis,biosynthesis of secondary metabolites,fatty acid degradation,and tyrosine metabolism.By up-regulating the gene encoding ethanol dehydrogenase,white lotus rapidly stimulates plant defense responses,then startups defensive system and elevates resistance to disease.Detoxification was achieved by upregulating the glutathione-transferase gene involved in the glutathione metabolic pathway.The cell wall of white lotus was modified to resist Fusarium oxysporum invasion by upregulating the genes of enzyme encoding flavonoids,lignin and hydroxycinnamamide.4.The full-length c DNA named Nn ACT3 and Nn ACT5 of the agmatine coumaroyl transferase gene were firstly cloned by RT-PCR and RACE from white lotus,and the open reading frame（ORF）all encoded 429 amino acids.PBWA（V）hs-osgfp-Nn ACT3 and p BWA（V）hs-osgfp-Nn ACT5 recombinant plasmids were obtained by homologous cloning.The overexpression recombinant plasmid was transferred to Colombian wild-type Arabidopsis thaliana via agrobacterium-mediated transfer.Finally,we acquired ten PBWA（V）hs-osgfp-Nn ACT3 and ten p BWA（V）hs-osgfp-Nn ACT5 overexpressed transgenic Arabidopsis thaliana.