| Molecular mechanisms of selective nodulation for rhizobia have long been the focus of the research and a key factor to limit their potential of nitrogen fixation.Based on our extensive study in the past,we have got some basic knowledge of the process of selective nodulation and impact factors,and identified certain genes associated with rhizobial symbiosis,nitrogen fixation and selective nodulation.However,we still lack the systematic understanding of the relationship between these strains' genomic,genetic regulation,differences in gene expression,and gene interactions with the host.This study took two Bradyrhizobium diazoefficiens strains that demonstrated completely different nodulation abilities,strain 4534being the most competitive and strain 4222 being the least competitive in nodulation as the research materials.With the approach of comparative transcriptomic,we compared the gene expression of these two B.diazoefficiens strains in response to soybean root exudates,in order to further explore the mechanisms of selective nodulation and the key impacting factors.The main results are as follows:1.The study designed and improved a preparation method to effectively promote soybean root exudates.One of the difficulties of rhizobial nod genes inducing test is how to get effective soybean rhizosphere secretion inducer,which is also the experimental basis for the following research.After repeated experiments,we designed an efficient preparation method of collecting soybean root exudates for selective nodulation,and select the influence of rhizosphere secretion to nod gene expression as an evaluation criterion.The comparative proteomic and transcriptomic results proved that this method is effective.2.In order to find the competitiveness-related genes in B.diazoefficiens,the transcriptome of twosoybean root exudates treated B.diazoefficiens with completely different nodulation abilities(B.diazoefficiens 4534 and B.diazoefficiens 4222)were sequenced and compared.With the treatment of soybean root exudates,the strain 4534 contains 8072 unigenes that can be annotated in the databases of NR,KEGQ Swiss-Prot,GO and COG database while the strain 4222 had 8233 unigenes.Those unigenes were mainly involved in biological process(i.g.,regulation,localization,metabolic process,regulation of biological progress and single-organism process),cell part(i.g.,cell,cell part and macromolecular complex in cellular component cluster)and molecular function(i.g.,bing and catalytic activity).3.With the treatment of soybean root exudates,the strain 4534 had 457 differential expressed genes,with 253 up-regulated and 204 down-regulated.The strain 4222 just had 293 differential expressed genes,with 108 up-regulated and 185 down-regulated.Analysis of enriched GO functional pathways,biological metabolic pathways,and enriched KEGG pathways showed that with the treatment of soybean rhizosphere secretion,the differences of genes related to flagellum synthesis,signal stimulation and tropism were significant in strain 4534,and genes related to flagellum synthesis were changed in the strain 4222.4.Compared with the strain 4222,more amounts and categories of differential expressed genes were detected in the strain 4534.Based on the analysis of differential genes,some differentially expressed genes are associated with a two-component system(i.g.,nodW,phyR-?EcfG),bacterial chemotaxis(i.g.,cheA,unigene04832),ABC transport proteins(i.g.,unigene02212),IAA(indole-3-acetic acid)metabolism(i.g.,nthA,nthB),and metabolic fitness(i.g.,put.),which may explain the higher nodulation competitivenessof B.diazoefficiens in the rhizosphere.Our results also showed that soybean rhizosphere secretion was involved not only in nodulation-related gene expression,but also in up regulation of metabolic fitness genes.Differentially expressed genes were monitored by quantitative real-time PCR,and 10 genes were found to be significantly up-regulated after encountering with B.diazoefficiens,which were consistent with the transcriptome analysis. |
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