Massively parallel sequencing holds great promise for expression profiling, as it combines the high throughput of SAGE with the accuracy of EST sequencing. Nevertheless, until now only very limited information had been available on the suitability of the current technology to meet the requirements. Here, we evaluate the potential of 454 sequencing technology for expression profiling using Drosophila melanogaster. We show that short (< ∼80 bp) and long (> ∼300-400 bp) cDNA fragments are under-represented in 454 sequence reads. Nevertheless, sequencing of 3Ј cDNA fragments generated by nebulization could be used to overcome the length bias of the 454 sequencing technology. Gene expression measurements generated by restriction analysis and nebulization for fragments within the 80-to 300-bp range showed correlations similar to those reported for replicated microarray experiments (0.83-0.91); 97% of the cDNA fragments could be unambiguously mapped to the genomic DNA, demonstrating the advantage of longer sequence reads. Our analyses suggest that the 454 technology has a large potential for expression profiling, and the high mapping accuracy indicates that it should be possible to compare expression profiles across species.[Supplemental material is available online at www.genome.org. The EST sequences have been deposited in GenBank under accession nos. EV574767-EV600806.]Gene expression technologies have greatly matured over the past years, but it has become clear that hybridization-based approaches have obvious limitations in cross-species comparisons (Gilad et al. 2005(Gilad et al. , 2006. Probably the most eminent problems are mismatches in heterologous probes and probe-specific hybridization kinetics, which complicate the design of speciesspecific oligonucleotide arrays. Alternatively, sequencing-based approaches could be used to measure gene expression if the sequence reads could be unambiguously mapped to the corresponding transcripts. While the short sequence reads of serial analysis of gene expression (SAGE) (Velculescu et al. 1995) and related techniques are severely limited by the requirement of a reliable genome annotation, the recently developed 454 sequencing technology (Margulies et al. 2005) may provide sufficient sequence information to overcome this limitation at moderate costs.In this study, we evaluate the potential of 454 sequencing technology to serve as a reliable tool for expression profiling. We show that 454 sequencing technology has a biased representation of cDNA fragments with different length. However, in combination with random breakage of the cDNAs by nebulization, 454 sequencing provides an excellent tool for expression profiling. The high accuracy with which we could map the sequenced fragments onto the Drosophila melanogaster genome suggests that 454 sequencing has great potential for interspecific expression profiling. Results Conceptual designMeasuring gene expression by sequencing requires only that a proportion of the transcript be analyzed. We sequenced a 3Ј region of the cDNA to...

Insects and their arthropod relatives including mites, spiders, and crustaceans play major roles in the world's terrestrial, aquatic, and marine ecosystems. Arthropods compete with humans for food and transmit devastating diseases. They also comprise the most diverse and successful branch of metazoan evolution, with millions of extant species. Here, we describe an international effort to guide arthropod genomic efforts, from species prioritization to methodology and informatics. The 5000 arthropod genomes initiative (i5K) community met formally in 2012 to discuss a roadmap for sequencing and analyzing 5000 high-priority arthropods and is continuing this effort via pilot projects, the development of standard operating procedures, and training of students and career scientists. With university, governmental, and industry support, the i5K Consortium aspires to deliver sequences and analytical tools for each of the arthropod branches and each of the species having beneficial and negative effects on humankind.

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We investigated the existence of species-specific associations between Brazilian coral species and bacteria. Pyrosequencing of the V3 region of the 16S rDNA was used to analyze the taxonomic composition of bacterial communities associated with the mucus of four coral species (Madracis decactis, Mussismilia hispida, Palythoa caribaeorum, and Tubastraea coccinea) in two seasons (winter and summer), which were compared with the surrounding water and sediment. The microbial communities found in samples of mucus, water, and sediment differed according to the composition and relative frequency of OTUs. The coral mucus community seemed to be more stable and resistant to seasonal variations, compared to the water and sediment communities. There was no influence of geographic location on the composition of the communities. The sediment community was extremely diverse and might act as a "seed bank" for the entire environment. Species-specific OTUs were found in P. caribaeorum, T. coccinea, and M. hispida.

Mitochondria play an essential role in several cellular processes. Nevertheless, very little is known about patterns of gene expression of genes encoded by the mitochondrial DNA (mtDNA). In this study, we used next-generation sequencing (NGS) for transcription profiling of genes encoded in the mitochondrial genome of Drosophila melanogaster and D. pseudoobscura. The analysis of males and females in both species indicated that the expression pattern was conserved between the two species, but differed significantly between both sexes. Interestingly, mRNA levels were not only different among genes encoded by separate transcription units, but also showed significant differences among genes located in the same transcription unit. Hence, mRNA abundance of genes encoded by mtDNA seems to be heavily modulated by post-transcriptional regulation. Finally, we also identified several transcripts with a noncanonical structure, suggesting that processing of mitochondrial transcripts may be more complex than previously assumed.