Project description:We investigated the anode-specific responses of Shewanella oneidensis MR-1, an exoelectroactive ammaproteobacterium, using for the first time iTRAQ and 2D-LC MS/MS driven membrane proteomics to compare protein abundances in S. oneidensis when generating power in MFCs, and growing in a continuous culture.
Project description:High-resolution tiling analysis of the MR-1 transcriptome under diverse growth conditions The conditions include aerobic growth in Luria-Bertani broth (LB), aerobic growth in defined lactate minimal medium, anaerobic growth in defined lactate minimal medium with 20mM dimethyl sulfoxide as the electron acceptor, anaerobic growth in defined lactate minimal medium with 10mM iron (III) citrate as the electron acceptor, 10 minutes post heat shock at 42oC see GSE39468 for tiling data on lactate minimal media
Project description:We combined high-resolution tiling microarrays and 5'-end RNA sequencing to obtain a genome-wide map of transcription start sites (TSSs) for Shewanella oneidensis MR-1. To test the reliability of these TSSs, we compared our result to those from differential RNA sequencing (dRNA-seq), which discriminates primary and processed ends of transcripts. We found that our identified TSSs tend to have significantly more mapped reads in the TEX(+) sample than the TEX(-) sample. Overall, the dRNA-seq results support the validity of our predictions for TSS.
Project description:Purpose: The goals of this study are to find out the differential expression genes in the fadR mutant strain(ΔfadR) compared with wild-type (WT) and to further explore the regulation mechanisms of fadR. Methods: Shewanella oneidensis MR-1 WT and ΔfadR were collected in log phage(OD~0.6). RNA extraction was performed using the RNeasy minikit (Qiagen) and the RNA was quantified by using a NanoVue spectrophotometer (GE Healthcare). RNA seq was performed using Illumina NextSeq 500, 2×150 bp. Results: Our study represents that the expression of 146 genes were decreased and 94 genes were increased inΔfadR compared with WT. Branched-chain keto acid dehydrogenase (BKD) produces corresponding branched-chain acyl coenzyme A which further participating branchend-chain fatty acids synthesis. The expression of bkdA2 was also promoted in △fadR compared with WT. Conclusions: Combined with our expression results, it declared that FadR can suppress bkd operon in some degree,which further increase the synthesis of branched-chain fatty acids in ΔfadR.
Project description:Shewanella oneidensis MR-1 is a platform microorganism for understanding extracellular electron transfer (EET) with a fully sequenced and annotated genome. In comparison to other model microorganisms such as Escherichia coli, the available plasmid parts (such as promoters and replicons) are not sufficient to conveniently and quickly fine-tune the expression of multiple genes in S. oneidensis MR-1. Here, we constructed and characterized a plasmid toolkit that contains a set of expression vectors with a combination of promoters, replicons, antibiotic resistance genes, and an RK2 origin of transfer (oriT) cassette, in which each element can be easily changed by fixed restriction enzyme sites. The expression cassette is also compatible with BioBrick synthetic biology standards. Using green fluorescent protein (GFP) as a reporter, we tested and quantified the strength of promoters. The copy number of different replicons was also measured by real-time quantitative PCR. We further transformed two compatible plasmids with different antibiotic resistance genes into the recombinant S. oneidensis MR-1, enabling control over the expression of two different fluorescent proteins. This plasmid toolkit was further used for overexpression of the MtrCAB porin-c-type cytochrome complex in the S. oneidensis ΔmtrA strain. Tungsten trioxide (WO3) reduction and microbial fuel cell (MFC) assays revealed that the EET efficiency was improved most significantly when MtrCAB was expressed at a moderate level, thus demonstrating the utility of the plasmid toolkit in the EET regulation in S. oneidensis. The plasmid toolkit developed in this study is useful for rapid and convenient fine-tuning of gene expression and enhances the ability to genetically manipulate S. oneidensis MR-1.