A microarray for assessing transcription from pelagic marine microbial taxa

Abstract

Metagenomic approaches have revealed unprecedented genetic diversity within microbial communities across vast expanses of the world's oceans. Linking this genetic diversity with key metabolic and cellular activities of microbial assemblages is a fundamental challenge. Here we report on a collaborative effort to design MicroTOOLs (Microbiological Targets for Ocean Observing Laboratories), a high-density oligonucleotide microarray that targets functional genes of diverse taxa in pelagic and coastal marine microbial communities. MicroTOOLs integrates nucleotide sequence information from disparate data types: genomes, PCR-amplicons, metagenomes, and metatranscriptomes. It targets 19 400 unique sequences over 145 different genes that are relevant to stress responses and microbial metabolism across the three domains of life and viruses. MicroTOOLs was used in a proof-of-concept experiment that compared the functional responses of microbial communities following Fe and P enrichments of surface water samples from the North Pacific Subtropical Gyre. We detected transcription of 68% of the gene targets across major taxonomic groups, and the pattern of transcription indicated relief from Fe limitation and transition to N limitation in some taxa. Prochlorococcus (eHLI), Synechococcus (sub-cluster 5.3) and Alphaproteobacteria SAR11 clade (HIMB59) showed the strongest responses to the Fe enrichment. In addition, members of uncharacterized lineages also responded. The MicroTOOLs microarray provides a robust tool for comprehensive characterization of major functional groups of microbes in the open ocean, and the design can be easily amended for specific environments and research questions.

A transcriptional cycle suited to daytime N2 fixation in the unicellular cyanobacterium Candidatus Atelocyanobacterium thalassa (UCYN-A)

“…From environmental RNA samples that contained UCYN-A, double-stranded (ds) cDNA was synthesized and amplified following the procedure described in Shilova et al. (54). Briefly, 400 ng RNA from each sample was used, and 1 μL of 1:100 dilution (corresponding to 4.7 attomoles of ERCC-0016) of the (External RNA Control Consortium, (55)) RNA spike-in mix 1 (Ambion®) was added before amplification to monitor the technical performance of the assay (55).…”

Techniques for Quantifying Phytoplankton Biodiversity

“…Based on this concept, arrays have now been designed to target specific genes in mixed communities. These array-based metagenomic techniques were first developed for soil microbial communities (e.g., GeoChip) (He et al 2007) but were then later designed for marine communities, including phytoplankton (Ottesen et al 2011, Rich et al 2008, Shilova et al 2014.…”

Complementary approaches to diagnosing marine diseases: a union of the modern and the classic

“…An increase in the reported frequency and severity of marine diseases [5,6] demands that complementary tools and approaches be used for rapid and effective diagnosis. Such tools are also critical to establish the essential baseline data (box 1) necessary for comparative investigations of marine epizootics [6,16,17]. Technologic approaches have recently advanced by leaps and bounds, providing exciting new diagnostic tools such as high-throughput sequencing, -omics (e.g.…”