401 research outputs found

    Simultaneous Determination of Various Isothiocyanates by RP-LC Following Precolumn Derivatization with Mercaptoethanol

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    Numerous isothiocyanates (ITCs) are poorly soluble in water which causes their precipitation in aqueous mobile phases used in reversed phase liquid chromatography (RP-LC), thus impacting the accuracy of the quantification. By comparing the amounts of ITCs injected and released from the column, losses could be estimated at 5–32% depending on polarities and concentrations. Results could be dramatically improved in terms of separation and quantification using RP-LC with a mercaptoethanol precolumn derivatization aimed at avoiding ITCs precipitation. The cancer chemoprotective allyl-ITC and sulforaphane were found in cabbage extracts at 1.2 and 2.7 μg g−1 fresh weight, respectively

    Methylation-capture and Next-Generation sequencing of free circulating DNA from human plasma

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    Background Free circulating DNA (fcDNA) has many potential clinical applications, due to the non-invasive way in which it is collected. However, because of the low concentration of fcDNA in blood, genome-wide analysis carries many technical challenges that must be overcome before fcDNA studies can reach their full potential. There are currently no definitive standards for fcDNA collection, processing and whole-genome sequencing. We report novel detailed methodology for the capture of high-quality methylated fcDNA, library preparation and downstream genome-wide Next-Generation Sequencing. We also describe the effects of sample storage, processing and scaling on fcDNA recovery and quality. Results Use of serum versus plasma, and storage of blood prior to separation resulted in genomic DNA contamination, likely due to leukocyte lysis. Methylated fcDNA fragments were isolated from 5 donors using a methyl-binding protein-based protocol and appear as a discrete band of ~180 bases. This discrete band allows minimal sample loss at the size restriction step in library preparation for Next-Generation Sequencing, allowing for high-quality sequencing from minimal amounts of fcDNA. Following sequencing, we obtained 37×106-86×106 unique mappable reads, representing more than 50% of total mappable reads. The methylation status of 9 genomic regions as determined by DNA capture and sequencing was independently validated by clonal bisulphite sequencing. Conclusions Our optimized methods provide high-quality methylated fcDNA suitable for whole-genome sequencing, and allow good library complexity and accurate sequencing, despite using less than half of the recommended minimum input DNA

    New genetic loci implicated in fasting glucose homeostasis and their impact on type 2 diabetes risk.

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    Levels of circulating glucose are tightly regulated. To identify new loci influencing glycemic traits, we performed meta-analyses of 21 genome-wide association studies informative for fasting glucose, fasting insulin and indices of beta-cell function (HOMA-B) and insulin resistance (HOMA-IR) in up to 46,186 nondiabetic participants. Follow-up of 25 loci in up to 76,558 additional subjects identified 16 loci associated with fasting glucose and HOMA-B and two loci associated with fasting insulin and HOMA-IR. These include nine loci newly associated with fasting glucose (in or near ADCY5, MADD, ADRA2A, CRY2, FADS1, GLIS3, SLC2A2, PROX1 and C2CD4B) and one influencing fasting insulin and HOMA-IR (near IGF1). We also demonstrated association of ADCY5, PROX1, GCK, GCKR and DGKB-TMEM195 with type 2 diabetes. Within these loci, likely biological candidate genes influence signal transduction, cell proliferation, development, glucose-sensing and circadian regulation. Our results demonstrate that genetic studies of glycemic traits can identify type 2 diabetes risk loci, as well as loci containing gene variants that are associated with a modest elevation in glucose levels but are not associated with overt diabetes

    Analysis of unresolved complex mixtures of hydrocarbons extracted from Late Archean sediments by comprehensive two-dimensional gas chromatography (GC×GC)

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    Author Posting. © Elsevier B.V., 2008. This is the author's version of the work. It is posted here by permission of Elsevier B.V. for personal use, not for redistribution. The definitive version was published in Organic Geochemistry 39 (2008): 846-867, doi:10.1016/j.orggeochem.2008.03.006.Hydrocarbon mixtures too complex to resolve by traditional capillary gas chromatrography display gas chromatograms with dramatically rising baselines or “humps” of coeluting compounds that are termed unresolved complex mixtures (UCMs). Because the constituents of UCMs are not ordinarily identified, a large amount of geochemical information is never explored. Gas chromatograms of saturated/unsaturated hydrocarbons extracted from Late Archean argillites and greywackes of the southern Abitibi Province of Ontario, Canada contain UCMs with different appearances or “topologies” relating to the intensity and retention time of the compounds comprising the UCMs. These topologies appear to have some level of stratigraphic organization, such that samples collected at any stratigraphic formation collectively are dominated by UCMs that either elute early- (within a window of C15-C20 of n-alkanes), early- to mid- (C15-C30 of n-alkanes), or have a broad UCM that extends through the entire retention time of the sample (from C15-C42 of n-alkanes). Comprehensive two-dimensional gas chromatography time-of-flight mass spectrometry (GC×GC-MS) was used to resolve the constituents forming these various UCMs. Early- to mid- eluting UCMs are dominated by configurational isomers of alkyl-substituted and non substituted polycyclic compounds that contain up to six rings. Late eluting UCMs are composed of C36-C40 mono-, bi-, and tricyclic archaeal isoprenoid diastereomers. Broad UCMs spanning the retention time of compound elution contain nearly the same compounds observed in the early-, mid-, and late retention time UCMs. Although the origin of the polycyclic compounds is unclear, the variations in the UCM topology appear to depend on the concentration of initial compound classes that have the potential to become isomerized. Isomerization of these constituents may have resulted from hydrothermal alteration of organic matter.This project was supported by NASA Exobiology grant #NAG5-13446 to Fabien Kenig. GC×GC analysis was supported by NSF grant IIS-0430835 and the Seaver Foundation to Christopher M. Reddy. Preparation of the archaeal biphytane standard was supported by NSF grant ARC-0520226 to Benjamin Van Mooy

    Bridging reproductive and microbial ecology: a case study in arbuscular mycorrhizal fungi

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    Offspring size is a key trait for understanding the reproductive ecology of species, yet studies addressing the ecological meaning of offspring size have so far been limited to macro-organisms. We consider this a missed opportunity in microbial ecology and provide what we believe is the first formal study of offspring-size variation in microbes using reproductive models developed for macro-organisms. We mapped the entire distribution of fungal spore size in the arbuscular mycorrhizal (AM) fungi (subphylum Glomeromycotina) and tested allometric expectations of this trait to offspring (spore) output and body size. Our results reveal a potential paradox in the reproductive ecology of AM fungi: while large spore-size variation is maintained through evolutionary time (independent of body size), increases in spore size trade off with spore output. That is, parental mycelia of large-spored species produce fewer spores and thus may have a fitness disadvantage compared to small-spored species. The persistence of the large-spore strategy, despite this apparent fitness disadvantage, suggests the existence of advantages to large-spored species that could manifest later in fungal life history. Thus, we consider that solving this paradox opens the door to fruitful future research establishing the relationship between offspring size and other AM life history traits

    When Your Decisions Are Not (Quite) Your Own: Action Observation Influences Free Choices

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    A growing number of studies have begun to assess how the actions of one individual are represented in an observer. Using a variant of an action observation paradigm, four experiments examined whether one person's behaviour can influence the subjective decisions and judgements of another. In Experiment 1, two observers sat adjacent to each other and took turns to freely select and reach to one of two locations. Results showed that participants were less likely to make a response to the same location as their partner. In three further experiments observers were asked to decide which of two familiar products they preferred or which of two faces were most attractive. Results showed that participants were less likely to choose the product or face occupying the location of their partner's previous reaching response. These findings suggest that action observation can influence a range of free choice preferences and decisions. Possible mechanisms through which this influence occurs are discussed

    Neighbour identity hardly affects litter-mixture effects on decomposition rates of New Zealand forest species.

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    The mass loss of litter mixtures is often different than expected based on the mass loss of the component species. We investigated if the identity of neighbour species affects these litter-mixing effects. To achieve this, we compared decomposition rates in monoculture and in all possible two-species combinations of eight tree species, widely differing in litter chemistry, set out in two contrasting New Zealand forest types. Litter from the mixed-species litter bags was separated into its component species, which allowed us to quantify the importance of litter-mixing effects and neighbour identity, relative to the effects of species identity, litter chemistry and litter incubation environment. Controlling factors on litter decomposition rate decreased in importance in the order: species identity (litter quality) >> forest type >> neighbour species. Species identity had the strongest influence on decomposition rate. Interspecific differences in initial litter lignin concentration explained a large proportion of the interspecific differences in litter decomposition rate. Litter mass loss was higher and litter-mixture effects were stronger on the younger, more fertile alluvial soils than on the older, less-fertile marine terrace soils. Litter-mixture effects only shifted percentage mass loss within the range of 1.5%. There was no evidence that certain litter mixtures consistently showed interactive effects. Contrary to common theory, adding a relatively fast-decomposing species generally slowed down the decomposition of the slower decomposing species in the mixture. This study shows that: (1) species identity, litter chemistry and forest type are quantitatively the most important drivers of litter decomposition in a New Zealand rain forest; (2) litter-mixture effects—although statistically significant—are far less important and hardly depend on the identity and the chemical characteristics of the neighbour species; (3) additive effects predominate in this ecosystem, so that mass dynamics of the mixtures can be predicted from the monocultures

    Three-Dimensional Geometric Analysis of Felid Limb Bone Allometry

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    Studies of bone allometry typically use simple measurements taken in a small number of locations per bone; often the midshaft diameter or joint surface area is compared to body mass or bone length. However, bones must fulfil multiple roles simultaneously with minimum cost to the animal while meeting the structural requirements imposed by behaviour and locomotion, and not exceeding its capacity for adaptation and repair. We use entire bone volumes from the forelimbs and hindlimbs of Felidae (cats) to investigate regional complexities in bone allometry.Computed tomographic (CT) images (16435 slices in 116 stacks) were made of 9 limb bones from each of 13 individuals of 9 feline species ranging in size from domestic cat (Felis catus) to tiger (Panthera tigris). Eleven geometric parameters were calculated for every CT slice and scaling exponents calculated at 5% increments along the entire length of each bone. Three-dimensional moments of inertia were calculated for each bone volume, and spherical radii were measured in the glenoid cavity, humeral head and femoral head. Allometry of the midshaft, moments of inertia and joint radii were determined. Allometry was highly variable and related to local bone function, with joint surfaces and muscle attachment sites generally showing stronger positive allometry than the midshaft.Examining whole bones revealed that bone allometry is strongly affected by regional variations in bone function, presumably through mechanical effects on bone modelling. Bone's phenotypic plasticity may be an advantage during rapid evolutionary divergence by allowing exploitation of the full size range that a morphotype can occupy. Felids show bone allometry rather than postural change across their size range, unlike similar-sized animals
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