In this study, the authors focused on the well-characterized surface-water microbial community of Hawaii Ocean Time Series station ALOHA in the North Pacific Subtropical Gyre. RNA-amplification methods were first adapted to enable large amounts of cDNA to be generated from small quantities of community RNA. This method involved reverse transcription of bacterial RNA to which poly-A tails had been attached, a technique that amplified the RNA signal by approximately 1,000-fold. To test the reliability of this amplification method, the authors calibrated their results against the predominant and well-characterized cyanobacteria genus Prochlorococcus. cDNA was then extracted from picoplankton field samples using this new amplification method, analysed by pyrosequencing and compared with community genomic DNA from the same sample. Last, the cDNA and genomic-DNA sequences obtained were compared with those compiled in the NCBI non-redundant protein database and to predicted peptides from the Global Ocean Sampling metagenomics dataset to allow functional interpretation of pyrosequencing transcript estimates.
Clear patterns were observed in community gene-expression distribution for specific taxa, genes and protein families. Consistent with previous studies, a large number of the detected genes were associated with key metabolic pathways, such as photosynthesis, nitrogen acquisition and carbon fixation, which indicates possible roles for these genes in various biogeochemical cycles. Most intriguingly, however, approximately 50% of the detected transcript sequences were unique and their functions unknown. Furthermore, hypothetical genes were highly expressed, again indicating that unknown proteins are functionally important.
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