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.
Project description:To address the question of how photosynthetic bacterium Rhodopseudomonas palustris differentially regulates gene expression of three nitrogenase isozymes (Mo, V, and Fe nitrogenases), we constructed Mo strain (Mo nitrogenase only strain), V strain (V nitrogenase only strain), and Fe strain (Fe nitrogenase only strain), and analyzed the whole genome transcriptome profiles of each mutant and wild-type cells grown under nitrogen-fixing conditions. Keywords: Genetic modification
Project description:The purpose of the study is to identify Irr-responsive genes in the bacterium Bradyrhizobium japonicum. Parent strain LO was compared to irr mutant strain LODTM5 by whole genome microarray analysis. Both cell types were grown in iron-limited media. Keywords: Comparison of B. japonicum wild type and mutant cells
Project description:Differences in genome size and gene content are among the most important signatures of microbial adaptation and genome evolution. Here, we investigated the patterns of genome variation among strains of the symbiotic nitrogen-fixing bacterium Sinorhizobium meliloti. Using the sequenced strain Rm1021 as a reference, the genome size and gene content variations were analyzed among ten diverse natural strains, through pulse field gel electrophoresis (PFGE) and whole-genome microarray hybridizations. Our PFGE analysis showed a genome size range of 6.45-7.01Mbp, with the greatest variation arising from the pSymA replicon, followed by that of pSymB. No observable size difference was evident among the chromosomes. Consistent with this pattern of size differences, 41.2% of ORFs on pSymA were variably absent/present, followed by 12.7% on pSymB, and 3.7% on the chromosome. However, the percentages of ORFs that were variably duplicated were more evenly distributed among the three replicons, 11.0%, 16.5% and 15.3% respectively for ORFs on pSymA, pSymB and the chromosome. Among the 10 strains, the percentages of absent ORFs ranged from 1.51% to 6.35% and those of duplicated ORFs ranged from 0.27% to 8.56%. Our analyses showed that host plants, geographic origins, multilocus enzyme electrophoretic types, and replicon sizes had little influence on the distribution patterns of absent or duplicated ORFs. The proportions of ORFs that were either variably absent/present or variably duplicated differed greatly among the functional categories, for each of the three replicons as well as for the whole genome. Interestingly, we observed positive correlations among the three replicons in their numbers of absent ORFs as well as the numbers of duplicated ORFs, consistent with coordinated gene gains/losses in this important bacterium in nature. microarray:Sm6kOligo
Project description:Mycorrhiza helper bacteria (MHB) promote the formation of ectomycorrhizae between tree roots and ectomycorrhizal fungi. Despite the high relevance of MHB for forestry and for sustainable tree production in tree nurseries, little is known about the properties of the bacteria that contribute to their helper abilities. The MHB strain Pseudomonas fluorescens BBc6R8 is used as a model to study the mechanisms of the helper effect. We took advantage of new technologies to obtain, for the first time, the whole genome sequence of an MHB. Analyses reveal an important plasticity of the genome with numerous functions acquired by horizontal gene tranfer. Genome mining was combined with transcriptomic and mutagenesis approaches to reveal molecular determinants of the helper effect. The data suggest that the production of helper molecules is likely to be constitutive in vitro. The helper effect appears to be pleiotropic and to rely, for a substantial part, on trophic interactions. Despite its helper abilities, the bacterium is also able in specific conditions to outcompete ectomycorrhizal fungi and inhibit their growth. We conclude that the helper bacterium possess a broad range of properties whose expression depending on the biotic and abiotic conditions can result in either a beneficial, neutral or antagonistic interaction between the plant, the ectomycorrhizal fungus and the bacterium.
Project description:The proteome of the anaerobic bacterium Dehalococcoides mccartyi strain CBDB1 from the phylum Chloroflexi was investigated. D. mccartyi strain CBDB1 is a model organism for organohalide respiration where halogenated organic compounds serve as terminal electron acceptors. A wide range of halogenated organic compounds have been shown to be dehalogenated by the strain CBDB1. Therefore, D. mccartyi strain CBDB1 is a promising candidate for bioremediation application. Proteomic analysis of cultures grown with hexachlorobenzene as only electron acceptor resulted in identification of 8,491 distinct peptides which represents 1,023 proteins. A coverage of 70% of the 1,458 annotated proteins for strain CBDB1 was achieved. In addition, a spectral library was created from the annotated spectra. By using proteogenomics, 18 previously not annotated peptides were identified which contribute to four proteins previously not annotated and corrections in length of eight protein coding sequences.