1,721,094 research outputs found
High-throughput screening for in planta characterization of VOC biosynthetic genes by PTR-ToF-MS
Full text linkFunctional characterization of plant volatile organic compound (VOC) biosynthetic genes and elucidation of the biological
function of their products often involve the screening of large numbers of plants from either independent transformation
events or mapping populations. The low time resolution of standard gas chromatographic methods, however, represents a
major bottleneck for in planta genetic characterization of VOC biosynthetic genes. Here we present a fast and highly-sensitive
method for the high-throughput characterization of VOC emission levels/patterns by coupling a Proton Transfer Reaction
Time-of-Flight Mass Spectrometer to an autosampler for automation of sample measurement. With this system more than
700 samples per day can be screened, detecting for each sample hundreds of spectrometric peaks in the m/z 15–300 range.
As a case study, we report the characterization of VOC emissions from 116 independent Arabidopsis thaliana lines transformed
with a putative isoprene synthase gene, confirming its function also when fused to a C-terminal 3×FLAG tag. We
demonstrate that the method is more reliable than conventional characterization of transgene expression for the identification
of the most highly isoprene-emitting lines. The throughput of this VOC screening method exceeds that of existing alternatives,
potentially allowing its application to reverse and forward genetic screenings of genes contributing to VOC emission,
constituting a powerful tool for the functional characterization of VOC biosynthetic genes and elucidation of the biological
functions of their products directly in planta
Chlorophyll binding to monomeric light-harvesting complex - A mutation analysis of chromophore-binding residues
No full textThe chromophore binding properties of the higher plant light-harvesting complex II have been studied by site-directed mutagenesis of pigment-binding residues. Mutant apoproteins were overexpressed in Escherichia coli and then refolded in vitro with purified chromophores to yield holoproteins selectively affected in chlorophyll-binding sites. Biochemical and spectroscopic characterization showed a specific loss of pigments and absorption spectral forms for each mutant, thus allowing identification of the chromophores bound to most of the binding sites. On these bases a map for the occupancy of individual sites by chlorophyll a and chlorophyll b is proposed. In some cases a single mutation led to the loss of more than one chromophore indicating that four chlorophylls and one xanthophyll could be bound by pigment-pigment interactions. Differential absorption spectroscopy allowed identification of the Q(y) transition energy level for each chlorophyll within the complex. It is shown that not only site selectivity is largely conserved between light-harvesting complex II and CP29 but also the distribution of absorption forms among different protein domains, suggesting conservation of energy transfer pathways within the protein and outward to neighbor subunits of the photosystem
Overexpression of isoprene synthase affects ABA-and drought-related gene expression and enhances tolerance to abiotic stress
Full text linkIsoprene is the most abundant single biogenic volatile compound emitted by plants. Despite the relevance of this molecule to plant abiotic resistance and its impact on global atmospheric chemistry, little is known about the details of its mechanism of action. Here, we characterized through both physiological and molecular methods the mechanisms of action of isoprene using model transgenic arabidopsis lines overexpressing a monocot isoprene synthase gene. Our results demonstrated the effect that isoprene had on ABA signaling at different tissue-specific, spatial, and temporal scales. In particular, we found that isoprene enhanced stomatal sensitivity to ABA through upregulation of RD29B signaling gene. By contrast, isoprene decreased sensitivity to ABA in germinating seeds and roots, suggesting tissue-specific mechanisms of action. In leaves, isoprene caused the downregulation of COR15A and P5CS genes, suggesting that the enhanced tolerance to water-deprivation stress observed in isoprene-emitting plants may be mediated chiefly by an enhanced membrane integrity and tolerance to osmotic stress
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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