A perspective: Metatranscriptomics as a tool for the discovery of novel biocatalysts

Warnecke, Falk; Hess, Matthias

doi:10.1016/j.jbiotec.2009.03.022

Cited by 90 publications

(46 citation statements)

References 33 publications

(47 reference statements)

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“…Extraction was followed by RNA purification using the RNAeasy minikit (Qiagen), including an on-column DNase I (Roche, Germany) treatment as described previously (52). The enrichment of mRNA was performed by the selective removal of 16S and 23S rRNA using oligonucleotide probes attached to magnetic beads according to the manufacturer's protocol (Microbexpress; Ambion, Applied Biosystems, Niewerkerk a/d Ijssel, the Netherlands) (44). Total RNA and enriched mRNA yields were quantified spectrophotometrically (NanoDrop 1000; Nanodrop Technologies, Wilmington, DE), and total RNA quality was assessed by a microfluidics-based electrophoresis system (Experion RNA StdSens; Bio-Rad Laboratories Inc.).…”

Section: Methodsmentioning

confidence: 99%

Comparative Analysis of Lactobacillus plantarum WCFS1 Transcriptomes by Using DNA Microarray and Next-Generation Sequencing Technologies

Leimena

Wels

Bongers³

et al. 2012

Appl Environ Microbiol

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ABSTRACTRNA sequencing is starting to compete with the use of DNA microarrays for transcription analysis in eukaryotes as well as in prokaryotes. The application of RNA sequencing in prokaryotes requires additional steps in the RNA preparation procedure to increase the relative abundance of mRNA and cannot employ the poly(T)-primed approach in cDNA synthesis. In this study, we aimed to validate the use of RNA sequencing (direct cDNA sequencing and 3′-untranslated region [UTR] sequencing) usingLactobacillus plantarumWCFS1 as a model organism, employing its established microarray platform as a reference. A limited effect of mRNA enrichment on genome-wide transcript quantification was observed, and comparative transcriptome analyses were performed forL. plantarumWCFS1 grown in two different laboratory media. Microarray analyses and both RNA sequencing methods resulted in similar depths of analysis and generated similar fold-change ratios of differentially expressed genes. The highest overall correlation was found between microarray and direct cDNA sequencing-derived transcriptomes, while the 3′-UTR sequencing-derived transcriptome appeared to deviate the most. Overall, a high similarity between patterns of transcript abundance and fold-change levels of differentially expressed genes was detected by all three methods, indicating that the biological conclusions drawn from the transcriptome data were consistent among the three technologies.

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Section: Methodsmentioning

confidence: 99%

Comparative Analysis of Lactobacillus plantarum WCFS1 Transcriptomes by Using DNA Microarray and Next-Generation Sequencing Technologies

Leimena

Wels

Bongers³

et al. 2012

Appl Environ Microbiol

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show abstract

“…However, the metagenomic approach does not identify the kinds of genes that are actually expressed in certain environments, which reflect the major functional activities and adaptations of the microorganisms that express them. Therefore, the metatranscriptomic approach has become increasingly important in defining these overall environmental gene expression profiles (7,50,61) and in identifying both the precursor and processing status of such transcripts. Therefore, a combination of metagenomic and metatranscriptomic approaches should allow an overview to be established of the diversity of microbial communities and the dynamics of their gene expression.…”

mentioning

confidence: 99%

Metatranscriptomic Analysis of Microbes in an Oceanfront Deep-Subsurface Hot Spring Reveals Novel Small RNAs and Type-Specific tRNA Degradation

Murakami

Fujishima

Tomita

et al. 2012

Appl Environ Microbiol

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Studies of small noncoding RNAs (sRNAs) have been conducted predominantly using culturable organisms, and the acquisition of further information about sRNAs from global environments containing uncultured organisms now is very important. In this study, hot spring water (57°C, pH 8.1) was collected directly from the underground environment at depths of 250 to 1,000 m in Yunohama, Japan, and small RNA sequences obtained from the environment were analyzed. A phylogenetic analysis of both archaeal and bacterial 16S rRNA gene sequences was conducted, and the results suggested the presence of unique species in the environment, corresponding to the Archaeal Richmond Mine Acidophilic Nanoorganisms (ARMAN) group and three new Betaproteobacteria. A metatranscriptomic analysis identified 64,194 (20,057 nonredundant) cDNA sequences. Of these cDNAs, 90% were either tRNAs, tRNA fragments, rRNAs, or rRNA fragments, whereas 2,181 reads (10%) were classified as previously uncharacterized putative candidate sRNAs. Among these, 15 were particularly abundant, 14 of which showed no sequence similarity to any known noncoding RNA, and at least six of which form very stable RNA secondary structures. The analysis of a large number of tRNA fragments suggested that unique relationships exist between the anticodons of the tRNAs and the sites of tRNA degradation. Previous bacterial tRNA degradation studies have been limited to specific organisms, such as Escherichia coli and Streptomyces coelicolor, and the current results suggest that specific tRNA decay occurs more frequently than previously expected.

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“…Because no primers or probes are required for direct sequencing, there is no need to anticipate important genes a priori and transcripts from· microbial assemblages are sequenced without bias. Furthermore, highly similar sequences, which might cross-hybridize on a microarray, can be distinguished by having a unique sequence (Warnecke and Hess, 2009;Shi et al, 2009). Experimental metatranscriptomics involves assessing changes in transcription in a particular environment and is a powerful tool for understanding the timing and regulation of complex microbial processes within communities and consortia, as well as microbial dexterity in response to changing conditions (Warnecke and Hess, 2009).…”

Section: Magneto-fishmentioning

confidence: 99%

“…Furthermore, highly similar sequences, which might cross-hybridize on a microarray, can be distinguished by having a unique sequence (Warnecke and Hess, 2009;Shi et al, 2009). Experimental metatranscriptomics involves assessing changes in transcription in a particular environment and is a powerful tool for understanding the timing and regulation of complex microbial processes within communities and consortia, as well as microbial dexterity in response to changing conditions (Warnecke and Hess, 2009). Community expression profiling via direct sequencing saves data obtained from individual metatranscriptomic studies and added to community databases.…”

Section: Magneto-fishmentioning

confidence: 99%

From Geocycles to Genomes and Back

Bose

Kopf

Newman

2014

Microbial Metal and Metalloid Metabolism

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A perspective: Metatranscriptomics as a tool for the discovery of novel biocatalysts

Cited by 90 publications

References 33 publications

Comparative Analysis of Lactobacillus plantarum WCFS1 Transcriptomes by Using DNA Microarray and Next-Generation Sequencing Technologies

Comparative Analysis of Lactobacillus plantarum WCFS1 Transcriptomes by Using DNA Microarray and Next-Generation Sequencing Technologies

Metatranscriptomic Analysis of Microbes in an Oceanfront Deep-Subsurface Hot Spring Reveals Novel Small RNAs and Type-Specific tRNA Degradation

From Geocycles to Genomes and Back

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