| Foxtail millet plays an important role in the production of coarse cereals,but in recent years,due to the large-scale occurrence of downy mildew,it has posed a serious threat to the yield and quality of foxtail millet.It’is difficult to culture the pathogen in vitro.At present,there are few studies on the cultivation,inoculation technology and the interaction mechanism between foxtail millet and downy mildew pathogen.In this study,we used the method of seed dressing of oospore and Jingu 21,a highly sensitive variety,to find out the most suitable infection condition of by Sclerospora graminicola measuring the morphological and physiological indexes of foxtail millet varity,and to explore the most suitable inoculation method of downy mildew by inoculating spores suspension in different growth stages and different parts of foxtail millet.Then,RNA-Seq technology was used to analyze the transcriptome of resistant / susceptible varieties at 12、 24 and 48 hours of seedling stage,to identify the differentially expressed genes,and to explore the candidate genes for disease resistance.The main results were as follows:1.The optimum infection temperature of Jingu 21 was determined by setting 5 different temperature treatments.The plant height,leaf length,leaf width and stem diameter were lower than those of the control group at the-18℃、4℃、15℃、 20℃、25℃,respectively.The morphological index,chlorophyll content and incidence rate of the tested foxtail millet were lower than those of the control groups.In conclusion,in the temperature range of-18℃~25℃,oospores can germinate and infect,the tempreture of 20℃ is the suitable for oospore germination.2.In order to find out the most suitable inoculation method for the downy mildew of foxtail millet,the seedlings were inoculated with resistant and susceptible foxtail millet varieties by immersing root and spraying immersing root at the seedling stage,heart injection at the booting stage,and ear injection at booting stage respectivly.By spraying root inoculation on the 5 d of resistant and susceptible varieties JG42 and JG21 in seedling stage,respectively.It was found that the susceptible variety JG21 became yellowish brown and withered and necrotic,and the morbidity and mortality were significantly higher than that of JG42.The survival rate of seedlings was the lowest(3%)when the immersing root time was 4 h.It was found that the leaves of JG21 was curled and loosely organized,and there were no obvious symptoms in the resistant variety.In the booting stage,hedgehog head was found in jingu 21,but there was no obvious symptom in resistant variety.It can be seen that spray seeding method at seedling stage and inoculation at booting stage are suitable for inoculation of S.graminicola.3.Spraying inoculation method was used to inoculate 12 h、24 h and 48 h respectively.Through the analysis of differential gene expression,Go enrichment and KEGG metabolic pathway,we found that MAPK cascade,WRKY transcription factors,genes encoding peroxidase,oxidoreductase,chitinase,serine/threonine protein kinase and cytochrome P450 were significantly induced;phenylpropane biosynthesis pathway and glutathione metabolism pathway,phenylalanine metabolism pathway and other related genes were differentially expressed in resistant varieties at different times of inoculation.It is speculated that the genes and important metabolic pathways differentially expressed in the resistant varieties may be involved in the regulation of resistance of foxtail millet to S.graminicola.4.Bioinformatics was used to analyze the basic characteristics,domain,collinearity and phylogenetic tree of the 28 differentially expressed WRKY transcription factors in the above transcriptome.The average number of amino acids was 349 AA,the theoretical isoelectric point of amino acids is between 5.26 and 11.49;domain analysis showed that 89.3% of transcription factors contain one WRKY domain;the 28 WRKY genes of foxtail millet and 15 resistant-related WRKY genes of rice are constructed into phylogenetic tree analysis,and the results showed that 9 pairs of genes are close to each other;the self collinearity analysis of foxtail millet showed that Seita.3g208900 and Seita.4g103800 are relatively close Strong collinearity indicates that it may have similar functions.The analysis also found that there were 24 WRKY collinearity genes between foxtail millet and green millet.In this study,the suitable temperature conditions for the infection of S.graminicola were clarified.The spray soaking method at seedling stage and at booting stage were initially suitable for Sclerospora graminicola infection.Using transcriptome sequencing,the differentially expressed genes in the resistant variety were identified,and the significantly enriched metabolic pathways and genes were screened out.Finally,the bioinformatics analysis of 28 WRKY transcription factors provided the theoretical basis for the identification,cloning and functional verification of resistant genes in foxtail millet. |
PDF Full Text Request