A custom oligonucleotide microarray analysis as a tool for dissecting soybean-bradyrhizobium japonicum nodule senescence

Abstract

[ACCESS RESTRICTED TO THE UNIVERSITY OF MISSOURI AT AUTHOR'S REQUEST.] Soybean root nodules are symbiotic organs housing Bradyrhizobium japonicum, a soil bacterium that reduces atmospheric nitrogen to ammonium. The bacterium receives carbon compounds from the plant, and the plant receives reduced nitrogen, the most limiting nutrient in agriculture for the synthesis of proteins, amino acids, nucleotides and vitamins. Most investigations on this symbiosis concern the formation of nodules whereas very few concern senescence. In this project, we aim to dissect the transcriptional networks that underlie nodule senescence employing oligonucleotide microarray technology, an obvious choice to identify global gene expression patterns during nodule development. The oligonucleotide (~70 mers) microarray contained 8,540 genes of the B. japonicum genome. The change in transcription during senescence was followed at multiple time points from 33- to 95- days after planting in the field. Bacteroids were extracted from soybean root nodules, mRNA was isolated, and cDNA synthesized in vitro and hybridized with B. japonicum oligonucleotide microarrays. Free-living B. japonicum 110 grown in defined media containing arabinose were used as reference design controls. A total of 5,488 genes were found to be differentially regulated. Cluster analysis generated several gene expression patterns. As expected, the genes for nitrogen fixation showed decreasing patterns during senescence. Surprisingly, a number of genes for flagella and chemotaxis showed increasing patterns as well genes for cell division and ATPase. Bacteroids were examined in-situ for the presence of flagella or pili. Flagella were not found, but structures resembling nanowires were found connecting the bacteroids. This study demonstrates how global analysis can lead to new and novel discoveries.--From public.pdf.