Project description:Single-cell RNA sequencing of Medicago truncatula roots was generated from nuclei using the 10X Genomics' Chromium technology. Roots were harvested and processed 24h after exposure to Ensifer meliloti (formerly Sinorhizobium meliloti).
Project description:Sinorhizobium meliloti can live as a soil saprophyte, and can engage in a nitrogen fixing symbiosis with plant roots. To succeed in such diverse environments, the bacteria must continually adjust gene expression. Transcriptional plasticity in eubacteria is often mediated by alternative sigma factors interacting with core RNA polymerase. The S. meliloti genome encodes 14 of these alternative sigmas, including two putative RpoH (heat shock) sigmas. We used custom Affymetrix Symbiosis Chips to characterize the global transcriptional response of S. meliloti rpoH1, rpoH2 and rpoH1 rpoH2 mutants during heat shock and stationary phase growth. Under these conditions, expression of over 300 genes is dependent on rpoH1 and rpoH2.
Project description:Sinorhizobium meliloti lives as a soil saprophyte, and engages in a nitrogen fixing symbiosis with plant roots. To succeed in such diverse environments, the bacteria must continually adjust gene expression. Transcriptional plasticity in eubacteria is often mediated by alternative sigma factors interacting with core RNA polymerase. The S. meliloti genome encodes 14 of these alternative sigmas, including 11 extracytoplasmic function (ECF) sigmas. We used custom Affymetrix Symbiosis Chips to characterize the global transcriptional response of S. meliloti overexpressing the ECF sigma factor, RpoE2. Our work identifies over 200 genes whose expression is dependent on RpoE2.
Project description:Many bacteria, often associated with eukaryotic hosts and of relevance for biotechnological applications, harbor a multipartite genome composed of more than one replicon. Biotechnologically relevant phenotypes are often encoded by genes residing on the secondary replicons. A synthetic biology approach to developing enhanced strains for biotechnological purposes could therefore involve merging pieces or entire replicons from multiple strains into a single genome. Here we report the creation of a genomic hybrid strain in a model multipartite genome species, the plant-symbiotic bacterium Sinorhizobium meliloti. We term this strain as cis-hybrid, since it is produced by genomic material coming from the same species' pangenome. In particular, we moved the secondary replicon pSymA (accounting for nearly 20% of total genome content) from a donor S. meliloti strain to an acceptor strain. The cis-hybrid strain was screened for a panel of complex phenotypes (carbon/nitrogen utilization phenotypes, intra- and extracellular metabolomes, symbiosis, and various microbiological tests). Additionally, metabolic network reconstruction and constraint-based modeling were employed for in silico prediction of metabolic flux reorganization. Phenotypes of the cis-hybrid strain were in good agreement with those of both parental strains. Interestingly, the symbiotic phenotype showed a marked cultivar-specific improvement with the cis-hybrid strains compared to both parental strains. These results provide a proof-of-principle for the feasibility of genome-wide replicon-based remodelling of bacterial strains for improved biotechnological applications in precision agriculture.
