DNA Methylation Modulatory Mechanism In Disease-resisitance Enhanced Maize Colonized With The Endophytic Bacillus Subtilis DZSY21

DNA methylation is a manifestation of epigenetic inheritance.It can not only play an important role in the growth and development of plants,but also have an important influence on the plant stress resistance and disease resistance.Previous study has been reported that maize treated with endophytic bacteria DZSY21 could obviously enhance the disease resistance to corn southern leaf blight.Based on the premise,we studied the molecular mechanism of DNA methylation in the process of enhancing maize disease resistance after endocytic DZSY21 treatment of maize.Firstly,the methylation-sensitive amplified polymorphism analysis(MSAP)was used to predict the genome-wide DNA methylation level of the maize treated with endophytic bacteria DZSY21 and the control maize genomic DNA,and found that the methylation level of genomic DNA of maize treated with endophytic bacteria decreased by 2.4% compared to genomic DNA of maize with no endophytic bacteria treated;Then,on the basis of this,the whole genome bisulfite sequencing(WGBS)was conducted to further research on the methylation of DZSY21-treated and control maize genome,and we has been known that there were three types of genomic DNA methylation,namely CG,CHG and CHH,methylation levels of all of them have been reduced,showing hypomethylation patterns by the analysis of the sequencing result;Statistical sequencing results were used to construct differentially methylated gene databases and screened log2Ratio(S/CK)values,141 genes with more significant differences were identified,of which,60.3% showed down-regulation,39.7% showed up-regulation,and down-regulated genes accounted for the majority.After that,PFam was searched to predict the protein domain function of the genes,then 16 genes related to disease resistance were screened,and then methylation specific PCR was carried out to verificate the change of DNA methylation of the 16 genes,then,10 suspected genes were got.DNA methylation has a negative effect on the transcription and expression of genes,gene expression would be increased while DNA methylation level was reduced,so fluorescence quantitative PCR was used to further verificate the difference of DNA methylation by the relative expression difference,and finally seven more distinct disease-resistance genes were obtained;At last,taking into account the changes in methylation and the relative expression levels and other factors,the gene GRM3 with distinctly different factors was selected,and c DNA was used as template to amplify the CDS sequence of the gene,and the p1301 a expression vector was constructed for the following functional identification.In summary,based on the findings in this study,we can deduce that the molecular mechanism of DNA methylation in disease-resistance enhanced maize colonizated with endophyte DZSY21 could be that endophyte DZSY21 can be colonizated in maize leaves,causing a decrease in DNA methylation of some disease resistance genes,and then leading to an increase in the expression level of these disease-resistant genes,thereby enhancing maize disease resistance.