Transcriptome Sequence And Analysis Of Biocontrol Bacillus Methylotrophicus NJ13 Induced By Ilyonectria Robusta

Bacillus methylotrophicus NJ13 is a biocontrol strain with good disease-preventing effect on Ilyonectria robusta.In this study,the interaction between NJ13 and Ilyonectria robusta DQX01 was carried out,and the key time points for inhibiting the spore germination of rust rot were determined.To understand the genes in NJ13 that may be involved in inhibiting spore germination of DQX01,the changes of differential expression gene and gene expression profile of NJ13 strain during the interaction were analyzed by transcriptome sequencing technology.The specific test results are as follows:1.The effect of NJ13 on the spore germination of DQX01 was observed in the laboratory.It was found that the spore germination rate of DQX01 treated with NJ13 was significantly lower than that of the clear water control group,and the spore morphology of DQX01 was correlated with the co-culture time of the two strains,The inhibition rate of DQX01 spore germination by NJ13 was more than 50%.Meanwhile,the spores gradually expand into the shape of a "rosy bead".The key time point of interaction between the two strains was determined to be 3 h after inoculation with NJ13.2.The transcriptome sequencing obtained a total of 13.73 G data volume.Definition |log2(FoldChange)|>5 is a differential expression gene,and a total of 51 genes is with highly differential expressed.Through being analysed the differential expressed genes,it was found that these differential genes were mainly enriched in molecular function of carbohydrate transport,ion transmembrane transport and so on,as well as carbohydrate metabolism and vitamin metabolism were involved in biological processes.At the same time,metabolic pathway analysis was performed on the obtained differential genes.3.From the highly expressed genes,6 genes were selected for real-time quantitative PCR analysis at different time points.The results showed that the gene interaction was highest at 3 h,with prolonging of time,the expressed value of the gene has a decreasing trend,indicating that the selection of the key time point of interaction 3 h is more accurate.Fluorescence quantification results showed that the up-regulated differentially expressed genes obtained by transcriptome sequencing were still up-regulated,and the down-regulated differentially express genes were still down-regulated.The detection results were almost the same as the transcriptome sequencing results,which indicated that the transcriptome sequencing results were reliable.4.From the highly expressed genes,primers of 36 different genes were selected and designed for cloning verification to verify the similarity between the target gene and the original differential gene,and to ensure the accuracy of the transcriptome sequencing gene.24 target genes were successfully verified by cloning,which indicated that the transcriptome sequencing was relatively reliable.