This article reviews basic concepts, general applications, and the potential impact of nextgeneration sequencing (NGS) technologies on genomics, with particular reference to currently available and possible future platforms and bioinformatics. NGS technologies have demonstrated the capacity to sequence DNA at unprecedented speed, thereby enabling previously unimaginable scientific achievements and novel biological applications. But, the massive data produced by NGS also presents a significant challenge for data storage, analyses, and management solutions. Advanced bioinformatic tools are essential for the successful application of NGS technology. As evidenced throughout this review, NGS technologies will have a striking impact on genomic research and the entire biological field. With its ability to tackle the unsolved challenges unconquered by previous genomic technologies, NGS is likely to unravel the complexity of the human genome in terms of genetic variations, some of which may be confined to susceptible loci for some common human conditions. The impact of NGS technologies on genomics will be far reaching and likely change the field for years to come.

The sweet melon fruit is characterized by a metabolic transition during its development that leads to extensive accumulation of the disaccharide sucrose in the mature fruit. While the biochemistry of the sugar metabolism pathway of the cucurbits has been well studied, a comprehensive analysis of the pathway at the transcriptional level allows for a global genomic view of sugar metabolism during fruit sink development. We identified 42 genes encoding the enzymatic reactions of the sugar metabolism pathway in melon. The expression pattern of the 42 genes during fruit development of the sweet melon cv Dulce was determined from a deep sequencing analysis performed by 454 pyrosequencing technology, comprising over 350,000 transcripts from four stages of developing melon fruit flesh, allowing for digital expression of the complete metabolic pathway. The results shed light on the transcriptional control of sugar metabolism in the developing sweet melon fruit, particularly the metabolic transition to sucrose accumulation, and point to a concerted metabolic transition that occurs during fruit development.

Chromatin modifications affect flowering time in the long-day plant Arabidopsis thaliana, but the role of histone methylation in flowering time regulation of rice (Oryza sativa), a short-day plant, remains to be elucidated. We identified a late-flowering long vegetative phase1 (lvp1) mutant in rice and used map-based cloning to reveal that lvp1 affects the SET domain group protein 724 (SDG724). SDG724 functions as a histone methyltransferase in vitro and contributes to a major fraction of global histone H3 lysine 36 (H3K36) methylation in vivo. Expression analyses of flowering time genes in wild-type and lvp1 mutants revealed that Early heading date1, but not Heading date1, are misregulated in lvp1 mutants. In addition, the double mutant of lvp1 with photoperiod sensitivity5 (se5) flowered later than the se5 single mutant, indicating that lvp1 delays flowering time irrespective of photoperiod. Chromatin immunoprecipitation assays showed that lvp1 had reduced levels of H3K36me2/3 at MADS50 and RFT1. This suggests that the divergent functions of paralogs RFT1 and Hd3a, and of MADS50 and MADS51, are in part due to differential H3K36me2/3 deposition, which also correlates with higher expression levels of MADS50 and RFT1 in flowering promotion in rice.

SummaryRaf®nose and stachyose are ubiquitous galactosyl-sucrose oligosaccharides in the plant kingdom which play major roles, second only to sucrose, in photoassimilate translocation and seed carbohydrate storage. These sugars are initially metabolised by a-galactosidases (a-gal). We report the cloning and functional expression of the ®rst genes, CmAGA1 and CmAGA2, encoding for plant a-gals with alkaline pH optima from melon fruit (Cucumis melo L.), a raf®nose and stachyose translocating species. The alkaline a-gal genes show very high sequence homology with a family of unde®ned`seed imbibition proteins' (SIPs) which are present in a wide range of plant families. In order to con®rm the function of SIP proteins, a representative SIP gene, from tomato, was expressed and shown to have alkaline a-gal activity. Phylogenetic analysis based on amino acid sequences shows that the family of alkaline a-gals shares little homology with the known prokaryotic and eukaryotic a-gals of glycosyl hydrolase families 27 and 36, with the exception of two cross-family conserved sequences containing aspartates which probably function in the catalytic step. This previously uncharacterised, plant-speci®c a-gal family of glycosyl hydrolases, with optimal activity at neutral-alkaline pH likely functions in key processes of galactosyl-oligosaccharide metabolism, such as during seed germination and translocation of RFO photosynthate.

Low temperature is one of the most severe environmental factors that impair plant growth and agricultural production. To investigate how Thellungiella halophila, an Arabidopsis-like extremophile, adapts to cold stress, a comparative proteomic approach based on two-dimensional electrophoresis was adopted to identify proteins that changed in abundance in Thellungiella rosette leaves during short term (6 h, 2 and 5 days) and long term (24 days) exposure to cold stress. Sixty-six protein spots exhibited significant change at least at one time point and maximal cold stress induced-proteome change was found in long-term cold stress group while the minimal change was found in 6-h cold treatment group. Fifty protein spots were identified by mass spectrometry analysis. The identified proteins mainly participate in photosynthesis, RNA metabolism, defense response, energy pathway, protein synthesis, folding and degradation, cell wall and cytoskeleton and signal transduction. These proteins might work cooperatively to establish a new homeostasis under cold stress. Nearly half of the identified cold-responsive proteins were associated with various aspects of chloroplast physiology suggesting that the cold stress tolerance of T. halophila is achieved, at least partly, by regulation of chloroplast function. All protein spots involved in RNA metabolism, defense response, protein synthesis, folding and degradation were found to be upregulated markedly by cold treatment, indicating enhanced RNA metabolism, defense and protein metabolism may play crucial roles in cold tolerance mechanism in T. halophila.

[1] Five new instruments for semicontinuous measurements of fine particle (PM2.5) nitrate and sulfate were deployed in the Atlanta Supersite Experiment during an intensive study in August 1999. The instruments measured bulk aerosol chemical composition at rates ranging from every 5 min to once per hour. The techniques included a filter sampling system with automated water extraction and online ion chromatographic (IC) analysis, two systems that directly collected particles into water for IC analysis, and two techniques that converted aerosol nitrate or sulfate either catalytically or by flash vaporization to gaseous products that were measured with gas analyzers. During the one-month study, 15-min integrated nitrate concentrations were low, ranging from about 0.1 to 3.5 mg m À3 with a mean value of 0.5 mg m À3 . Ten-minute integrated sulfate concentrations varied between 0.3 and 40 mg m À3 with a mean of 14 mg m À3 . By the end of the one-month study most instruments were in close agreement, with r-squared values between instrument pairs typically ranging from 0.7 to 0.94. Based on comparison between individual semicontinuous devices and 24-hour integrated filter measurements, most instruments were within 20-30% for nitrate ($0.1-0.2 mg m À3 ) and 10-15% for sulfate (1-2 mg m À3 ). Within 95% confidence intervals, linear regression fits suggest that no biases existed between the semicontinuous techniques and the 24-hour integrated filter measurements of nitrate and sulfate;, however, for nitrate, the semicontinuous intercomparisons showed significantly less variability than intercomparisons amongst the 24-hour integrated filters.

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A new fluoropolymer tube is proposed as the basis of a novel class of liquid core waveguide-based luminescence detectors. Both chemiluminescence and photoluminescence detectors are possible. In the latter case, illumination is transverse to the main axis of the tube. With such a geometry, it is even possible to operate without monochromators, although limits of detection do improve with the incorporation of monochromators. The nature of the design is such that it is particularly simple to fabricate detectors in a flow-through configuration and where the light from the cell is coupled to a photodetector by an optical fiber. No focusing optics are necessary. A number of applications are illustrated. Attainable limits (LODs, S/N = 3) of detection include 150 pM fluorescein with a 254-nm excitation source, 200 amol of fluorescein in a capillary electrophoresis setup with excitation by two blue light-emitting diodes, 35 nM NH(3) as the isoindole derivative in a flow injection analysis system using a photodiode detector, 50 nM methylene blue and 1 nM Rhodamine 560 using respectively red and green LED arrays and an avalanche photodiode and a PMT in a FIA configuration, 100 parts per trillion by volume gaseous formaldehyde as the Hantzsch reaction product with cyclohexanedione using a diffusion scrubber, 2.7 μM and 17 nM hypochlorite based on its chemiluminescence reaction with luminol with photodiode and PMT detectors, respectively, and 1 ppm SO(4)(2)(-) based on nephelometric detection at 470 nm. The approach described herein leads to particularly simple and inexpensive luminescence detectors with excellent sensitivity.