Project description:Sequencing the metatranscriptome can provide information about the response of organisms to varying environmental conditions. We present a methodology for obtaining random whole-community mRNA from a complex microbial assemblage using Pyrosequencing. The metatranscriptome had, with minimum contamination by ribosomal RNA, significant coverage of abundant transcripts, and included significantly more potentially novel proteins than in the metagenome. Keywords: metatranscriptome, mesocosm, ocean acidification
Project description:Metagenome sequencing enables discovery and genetic characterization of complex microbial communities from diverse ecosystems. However, determining the activity of isolates within a community using transcriptomics presents several challenges including the wide dynamic range of organismal and gene expression abundances, the presence of host RNA, and low microbial biomass at many body sites. To address these limitations, we developed “Targeted Expression Analysis Sequencing” or TEAL-seq. Targeting strategies enabled sensitive species-specific analyses of gene expression using highly multiplexed custom probe pools targeting about 1700 core and accessory genes of Staphylococcus aureus (S.a.) and S. epidermidis (S.e.), two key species of the skin microbiome. Two targeting methods were applied to mixed cultures and nasal swab specimens from human research participants. Both methods showed a high degree of specificity, with >90% reads on target, even in the presence of complex microbial or human background DNA/RNA. Targeting using molecular inversion probes demonstrated excellent correlation in inferred expression levels with bulk RNA-seq. Further, we show that a linear pre-amplification step to increase the amount of input nucleic acids for analysis was quite reproducible . While pre-amplification introduced some noise compared to non-amplified samples, it also enabled profiling of expression from as little as 1 ng of total RNA. TEAL-seq is much less expensive than bulk metatranscriptomic profiling and enables detection across a greater dynamic range. Custom probe pools are readily configurable and this strategy is broadly applicable for determining the transcriptional status of organisms in any microbial community.
Project description:The fate of the carbon stocked in permafrost soils following global warming and permafrost thaw is of major concern in view of the potential for increased CH4 and CO2 emissions from these soils. Complex carbon compound degradation and greenhouse gas emissions are due to soil microbial communities, but their composition and functional potential in permafrost soils are largely unknown. Here, a 2 m deep permafrost and its overlying active layer soil were subjected to metagenome sequencing, quantitative PCR, and microarray analyses. The active layer soil and 2 m permafrost soil microbial community structures were very similar, with Actinobacteria being the dominant phylum. The two soils also possessed a highly similar spectrum of functional genes, especially when compared to other already published metagenomes. Key genes related to methane generation, methane oxidation and organic matter degradation were highly diverse for both soils in the metagenomic libraries and some (e.g. pmoA) showed relatively high abundance in qPCR assays. Genes related to nitrogen fixation and ammonia oxidation, which could have important roles following climatic change in these nitrogen-limited environments, showed low diversity but high abundance. The 2 m permafrost soil showed lower abundance and diversity for all the assessed genes and taxa. Experimental biases were also evaluated and showed that the whole community genome amplification technique used caused large representational biases in the metagenomic libraries. This study described for the first time the detailed functional potential of permafrost-affected soils and detected several genes and microorganisms that could have crucial importance following permafrost thaw. A 2m deep permafrost sample and it overlying active layer were sampled and their metagenome analysed. For microarray analyses, 8 other soil samples from the same region were used for comparison purposes.
Project description:This data set contains 1376 mass spectrometry reads from root, rhizosphere and leaf sample of Populus Trichocarpa, as well as associated controls. This metabolomics data set was collected as part of a larger campaign which complements the metabolomics data with metagenome sequencing, transcriptomics, and soil measurement data.
Project description:Background: The soil environment is responsible for sustaining most terrestrial plant life on earth, yet we know surprisingly little about the important functions carried out by diverse microbial communities in soil. Soil microbes that inhabit the channels of decaying root systems, the detritusphere, are likely to be essential for plant growth and health, as these channels are the preferred locations of new root growth. Understanding the microbial metagenome of the detritusphere and how it responds to agricultural management such as crop rotations and soil tillage will be vital for improving global food production. Methods: The rhizosphere soils of wheat and chickpea growing under + and - decaying root were collected for metagenomics sequencing. A gene catalogue was established by de novo assembling metagenomic sequencing. Genes abundance was compared between bulk soil and rhizosphere soils under different treatments. Conclusions: The study describes the diversity and functional capacity of a high-quality soil microbial metagenome. The results demonstrate the contribution of the microbiome from decaying root in determining the metagenome of developing root systems, which is fundamental to plant growth, since roots preferentially inhabit previous root channels. Modifications in root microbial function through soil management, can ultimately govern plant health, productivity and food security.