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1.
Background
Non-invasive and high-throughput monitoring of drought in plants from its initiation to visible symptoms is essential to quest drought tolerant varieties. Among the existing methods, chlorophyll a fluorescence (ChlF) imaging has the potential to probe systematic changes in photosynthetic reactions; however, prerequisite of dark-adaptation limits its use for high-throughput screening.Results
To improve the throughput monitoring of plants, we have exploited their light-adaptive strategy, and investigated possibilities of measuring ChlF transients under low ambient irradiance. We found that the ChlF transients and associated parameters of two contrasting Arabidopsis thaliana accessions, Rsch and Co, give almost similar information, when measured either after ~20 min dark-adaptation or in the presence of half of the adaptive growth-irradiance. The fluorescence parameters, effective quantum yield of PSII photochemistry (ΦPSII) and fluorescence decrease ratio (R FD) resulting from this approach enabled us to differentiate accessions that is often not possible by well-established dark-adapted fluorescence parameter maximum quantum efficiency of PSII photochemistry (F V/F M). Further, we screened ChlF transients in rosettes of well-watered and drought-stressed six A. thaliana accessions, under half of the adaptive growth-irradiance, without any prior dark-adaptation. Relative water content (RWC) in leaves was also assayed and compared to the ChlF parameters. As expected, the RWC was significantly different in drought-stressed from that in well-watered plants in all the six investigated accessions on day-10 of induced drought; the maximum reduction in the RWC was obtained for Rsch (16%), whereas the minimum reduction was for Co (~7%). Drought induced changes were reflected in several features of ChlF transients; combinatorial images obtained from pattern recognition algorithms, trained on pixels of image sequence, improved the contrast among drought-stressed accessions, and the derived images were well-correlated with their RWC.Conclusions
We demonstrate here that ChlF transients and associated parameters measured even in the presence of low ambient irradiance preserved its features comparable to that of measured after dark-adaptation and discriminated the accessions having differential geographical origin; further, in combination with combinatorial image analysis tools, these data may be readily employed for early sensing and mapping effects of drought on plant’s physiology via easy and fully non-invasive means.2.
Background
To identify plant genes involved in various key traits, QTL mapping is a powerful approach. This approach is based on the use of mapped molecular markers to identify genomic regions controlling quantitative traits followed by a fine mapping and eventually positional cloning of candidate genes. Mapping technologies using SNP markers are still rather expensive and not feasible in every laboratory. In contrast, microsatellite (also called SSR for Simple Sequence Repeat) markers are technologically less demanding and less costly for any laboratory interested in genetic mapping.Results
In this study, we present the development and the characterization of a panel of 96 highly polymorphic SSR markers along the Arabidopsis thaliana genome allowing QTL mapping among accessions of the Versailles 24 core collection that covers a high percentage of the A. thaliana genetic diversity. These markers can be used for any QTL mapping analysis involving any of these accessions. We optimized the use of these markers in order to reveal polymorphism using standard PCR conditions and agarose gel electrophoresis. In addition, we showed that the use of only three of these markers allows differentiating all 24 accessions which makes this set of markers a powerful tool to control accession identity or any cross between any of these accessions.Conclusion
The set of SSR markers developed in this study provides a simple and efficient tool for any laboratory focusing on QTL mapping in A. thaliana and a simple means to control seed stock or crosses between accessions. 相似文献3.
Background
Cowpea (Vigna unguiculata L.) is an important grain and forage legume grown throughout sub-Saharan Africa primarily by subsistence farmers on poor, drought prone soils. Genetic improvement of the crop is being actively pursued and numerous functional genomics studies are underway aimed at characterizing gene controlling key agronomic characteristics for disease and pest resistances. Unfortunately, similar to other legumes, efficient plant transformation technology is a rate-limiting step in analysis of gene function in cowpea.Results
Here we describe an optimized protocol for the rapid generation of transformed hairy roots on ex vitro composite plants of cowpea using Agrobacterium rhizogenes. We further demonstrate the applicability of cowpea composite plants to study gene expression involved in the resistance response of the plant roots to attack by the root parasitic weed, Striga gesnerioides. The utility of the new system and critical parameters of the method are described and discussed herein.Conclusions
Cowpea composite plants offer a rapid alternative to methods requiring stable transformation and whole plant regeneration for studying gene expression in resistance or susceptibility responses to parasitic weeds. Their use can likely be readily adapted to look at the effects of both ectopic gene overexpression as well as gene knockdown of root associated defense responses and to the study of a broader range of root associated physiological and aphysiological processes including root growth and differentiation as well as interactions with other root pests, parasites, and symbionts. 相似文献4.
Background
The success of the microarray reproducibility is dependent upon the performance of standardized procedures. Since the introduction of microarray technology for the analysis of global gene expression, reproducibility of results among different laboratories has been a major problem. Two of the main contributors to this variability are the use of different microarray platforms and different laboratory practices. In this paper, we address the latter question in terms of how variation in one of the steps of a labelling procedure affects the cDNA product prior to microarray hybridization.Results
We used a standard procedure to label cDNA for microarray hybridization and employed different types of column chromatography for cDNA purification. After purifying labelled cDNA, we used the Agilent 2100 Bioanalyzer and agarose gel electrophoresis to assess the quality of the labelled cDNA before its hybridization onto a microarray platform. There were major differences in the cDNA profile (i.e. cDNA fragment lengths and abundance) as a result of using four different columns for purification. In addition, different columns have different efficiencies to remove rRNA contamination. This study indicates that the appropriate column to use in this type of protocol has to be experimentally determined. Finally, we present new evidence establishing the importance of testing the method of purification used during an indirect labelling procedure. Our results confirm the importance of assessing the quality of the sample in the labelling procedure prior to hybridization onto a microarray platform.Conclusion
Standardization of column purification systems to be used in labelling procedures will improve the reproducibility of microarray results among different laboratories. In addition, implementation of a quality control check point of the labelled samples prior to microarray hybridization will prevent hybridizing a poor quality sample to expensive micorarrays. 相似文献5.
Background
A plethora of concurrent cellular activities is mobilised in the adaptation of plants to adverse environmental conditions. This response can be quantified by physiological experiments or metabolic profiling. The intention of this work is to reduce the number of metabolic processes studied to a minimum of relevant parameters with a maximum yield of information. Therefore, we inspected 'summary parameters' characteristic for whole classes of antioxidative metabolites and key enzymes.Results
Three bioluminescence assays are presented. A horseradish peroxidase-based total antioxidative capacity (TAC) assay is used to probe low molecular weight antioxidants. Peroxidases are quantified by their luminol converting activity (LUPO). Finally, we quantify high molecular weight superoxide anion scavenging activity (SOSA) using coelenterazine. Experiments with Lepidium sativum L. show how salt, drought, cold, and heat influence the antioxidative system represented here by TAC, LUPO, SOSA, catalase, and glutathione reductase (GR). LUPO and SOSA run anti-parallel under all investigated stress conditions suggesting shifts in antioxidative functions rather than formation of antioxidative power. TAC runs in parallel with GR. This indicates that a majority of low molecular weight antioxidants in plants is represented by glutathione.Conclusion
The set of assays presented here is capable of characterising antioxidative activities in plants. It is inexpensive, quick and reproducible and delivers quantitative data. 'Summary parameters' like TAC, LUPO, and SOSA are quantitative traits which may be promising for implementation in high-throughput screening for robustness of novel mutants, transgenics, or breeds. 相似文献6.
Zachary G. Loman William V. Deluca Daniel J. Harrison Cynthia S. Loftin Brian W. Rolek Petra B. Wood 《Landscape Ecology》2018,33(1):77-91
Context
Species-specific models of landscape capability (LC) can inform landscape conservation design. Landscape capability is “the ability of the landscape to provide the environment […] and the local resources […] needed for survival and reproduction […] in sufficient quantity, quality and accessibility to meet the life history requirements of individuals and local populations.” Landscape capability incorporates species’ life histories, ecologies, and distributions to model habitat for current and future landscapes and climates as a proactive strategy for conservation planning.Objectives
We tested the ability of a set of LC models to explain variation in point occupancy and abundance for seven bird species representative of spruce-fir, mixed conifer-hardwood, and riparian and wooded wetland macrohabitats.Methods
We compiled point count data sets used for biological inventory, species monitoring, and field studies across the northeastern United States to create an independent validation data set. Our validation explicitly accounted for underestimation in validation data using joint distance and time removal sampling.Results
Blackpoll warbler (Setophaga striata), wood thrush (Hylocichla mustelina), and Louisiana (Parkesia motacilla) and northern waterthrush (P. noveboracensis) models were validated as predicting variation in abundance, although this varied from not biologically meaningful (1%) to strongly meaningful (59%). We verified all seven species models [including ovenbird (Seiurus aurocapilla), blackburnian (Setophaga fusca) and cerulean warbler (Setophaga cerulea)], as all were positively related to occupancy data.Conclusions
LC models represent a useful tool for conservation planning owing to their predictive ability over a regional extent. As improved remote-sensed data become available, LC layers are updated, which will improve predictions.7.
E. L. Loudermilk R. M. Scheller P. J. Weisberg Alec Kretchun 《Landscape Ecology》2017,32(7):1461-1472
Context
Forest landscapes are increasingly managed for fire resilience, particularly in the western US which has recently experienced drought and widespread, high-severity wildfires. Fuel reduction treatments have been effective where fires coincide with treated areas. Fuel treatments also have the potential to reduce drought-mortality if tree density is uncharacteristically high, and to increase long-term carbon storage by reducing high-severity fire probability.Objective
Assess whether fuel treatments reduce fire intensity and spread and increase carbon storage under climate change.Methods
We used a simulation modeling approach that couples a landscape model of forest disturbance and succession with an ecosystem model of carbon dynamics (Century), to quantify the interacting effects of climate change, fuel treatments and wildfire for carbon storage potential in a mixed-conifer forest in the western USA.Results
Our results suggest that fuel treatments have the potential to ‘bend the C curve’, maintaining carbon resilience despite climate change and climate-related changes to the fire regime. Simulated fuel treatments resulted in reduced fire spread and severity. There was partial compensation of C lost during fuel treatments with increased growth of residual stock due to greater available soil water, as well as a shift in species composition to more drought- and fire-tolerant Pinus jeffreyi at the expense of shade-tolerant, fire-susceptible Abies concolor.Conclusions
Forest resilience to global change can be achieved through management that reduces drought stress and supports the establishment and dominance of tree species that are more fire- and drought-resistant, however, achieving a net C gain from fuel treatments may take decades.8.
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10.
Jing Wang Chongnan Wang Yan Long Clare Hopkins Smita Kurup Kede Liu Graham J King Jinling Meng 《Plant methods》2011,7(1):1-7
Background
Protein extraction is a frequent procedure in biological research. For preparation of plant cell extracts, plant materials usually have to be ground and homogenized to physically break the robust cell wall, but this step is laborious and time-consuming when a large number of samples are handled at once.Results
We developed a chemical method for lysing Arabidopsis cells without grinding. In this method, plants are boiled for just 10 minutes in a solution containing a Ca2+ chelator and detergent. Cell extracts prepared by this method were suitable for SDS-PAGE and immunoblot analysis. This method was also applicable to genomic DNA extraction for PCR analysis. Our method was applied to many other plant species, and worked well for some of them.Conclusions
Our method is rapid and economical, and allows many samples to be prepared simultaneously for protein analysis. Our method is useful not only for Arabidopsis research but also research on certain other species. 相似文献11.
Context
Mosquito-borne diseases in the United States are notorious for their seemingly erratic temporal and spatial outbreak patterns in humans. Identifying linkages between fine-scale phenomenon and macroscale outbreak patterns can improve the understanding of these complex socio–ecological systems.Objectives
Using West Nile virus (WNV) as a model system, we examine whether fine-scale wetland characteristics—wetland size, connectivity, and inundation regime—have an important effect on macroscale human disease outbreak in the northeastern and midwestern United States.Methods
We take a spatially explicit GIS-based approach, derive new macroscale wetland metrics based on fine-scale wetland characteristics, and use county-level annual human WNV incidence data over 11 years to evaluate linkages between the fine and macroscales (1.6 million km2).Results
In regions dominated by the Culex tarsalis WNV vector, U.S. counties with a small average wetland size had more than 100 % higher human WNV incidence than counties with a large average wetland size. In regions dominated by the Cx. pipiens and Cx. restuans vectors, a low proportional area of connected wetland was associated with at least 50 % higher human WNV incidence than a high proportional area of connected wetland. Finally, Cx. tarsalis-dominated counties with a high proportional area of semi-permanent wetland that were experiencing drought conditions had over 300 % higher annual WNV incidence than drought-affected counties with a low proportional area of semi-permanent wetland.Conclusions
Our results suggest that phenomena occurring at the individual wetland scale may aggregate to influence macroscale human WNV outbreak patterns and may be mediated by the interplay of other factors such as vector species-specific traits and climate.12.
Background
In vivo detection of protein-bound genomic regions can be achieved by combining chromatin-immunoprecipitation with next-generation sequencing technology (ChIP-seq). The large amount of sequence data produced by this method needs to be analyzed in a statistically proper and computationally efficient manner. The generation of high copy numbers of DNA fragments as an artifact of the PCR step in ChIP-seq is an important source of bias of this methodology.Results
We present here an R package for the statistical analysis of ChIP-seq experiments. Taking the average size of DNA fragments subjected to sequencing into account, the software calculates single-nucleotide read-enrichment values. After normalization, sample and control are compared using a test based on the ratio test or the Poisson distribution. Test statistic thresholds to control the false discovery rate are obtained through random permutations. Computational efficiency is achieved by implementing the most time-consuming functions in C++ and integrating these in the R package. An analysis of simulated and experimental ChIP-seq data is presented to demonstrate the robustness of our method against PCR-artefacts and its adequate control of the error rate.Conclusions
The software ChIP-seq Analysis in R (CSAR) enables fast and accurate detection of protein-bound genomic regions through the analysis of ChIP-seq experiments. Compared to existing methods, we found that our package shows greater robustness against PCR-artefacts and better control of the error rate. 相似文献13.
Method: low-cost delivery of the cotton leaf crumple virus-induced gene silencing system 总被引:1,自引:0,他引:1
Background
We previously developed a virus-induced gene silencing (VIGS) vector for cotton from the bipartite geminivirusCotton leaf crumple virus (CLCrV). The original CLCrV VIGS vector was designed for biolistic delivery by a gene gun. This prerequisite limited the use of the system to labs with access to biolistic equipment. Here we describe the adaptation of this system for delivery by Agrobacterium (Agrobacterium tumefaciens). We also describe the construction of two low-cost particle inflow guns.Results
The biolistic CLCrV vector was transferred into two Agrobacterium binary plasmids. Agroinoculation of the binary plasmids into cotton resulted in silencing and GFP expression comparable to the biolistic vector. Two homemade low-cost gene guns were used to successfully inoculate cotton (G. hirsutum) and N. benthamiana with either the CLCrV VIGS vector or the Tomato golden mosaic virus (TGMV) VIGS vector respectively.Conclusions
These innovations extend the versatility of CLCrV-based VIGS for analyzing gene function in cotton. The two low-cost gene guns make VIGS experiments affordable for both research and teaching labs by providing a working alternative to expensive commercial gene guns. 相似文献14.
15.
Grant S. Joseph Colleen L. Seymour Bernard W. T. Coetzee Mduduzi Ndlovu Luana Deng Kelly Fowler James Hagan Brian J. Brooks Jackson A. Seminara Stefan H. Foord 《Landscape Ecology》2018,33(5):731-742
Context
With global change, microclimates become important refuges for temperature-sensitive, range-restricted organisms. In African savannas, woody vegetation on Macrotermes mounds create widely-dispersed microclimates significantly cooler than the surrounding matrix, which buffer against elevated temperatures at the finer scale of mounds, allowing species to persist at the landscape scale. Termite colonies cultivate symbiotic fungi to digest lignin, but the fungi require temperatures between 29 and 32 °C, which termites strive to maintain. Mound-associated vegetation is a hot-spot for elephant herbivory, so removal of woody species cover by elephants could influence mound-associated microclimates, impacting temperature regulation by termites.Objectives
We explored the interaction between two prominent ecosystem engineers (termites and elephants) to ascertain whether elephant removal of mound woody cover affects (1) external mound-associated microclimate and (2) internal mound temperature.Methods
We surveyed 44 mounds from three sites in Kruger National Park, South Africa, during an El Niño/Southern Oscillation-induced drought and heatwave, recording whether sub-canopy, external, mound-surface and internal mound temperatures varied with vegetation removal by elephant.Results
Elephant damage to mound-associated vegetation reduces the fine-scale microclimate effect provided by vegetation on Macrotermes mounds. Despite this, termites were able to regulate internal mound temperatures, whereas internal temperatures of abandoned mounds increased with elevated surface temperatures.Conclusions
Termites can persist despite loss of mound-associated microclimates, but the loss likely increases energetic costs of mound thermoregulation. Since mound vegetation buffers against drought, loss of widely-dispersed, fine-scale microclimates could increase as megaherbivores remain constrained to protected areas, impacting climate-sensitive organisms and ecosystem function at a range of scales.16.
Background
Histochemical staining of plant tissues with 4-dimethylaminocinnamaldehyde (DMACA) or vanillin-HCl is widely used to characterize spatial patterns of proanthocyanidin accumulation in plant tissues. These methods are limited in their ability to allow high-resolution imaging of proanthocyanidin deposits.Results
Tissue embedding techniques were used in combination with DMACA staining to analyze the accumulation of proanthocyanidins in Lotus corniculatus (L.) and Trifolium repens (L.) tissues. Embedding of plant tissues in LR White or paraffin matrices, with or without DMACA staining, preserved the physical integrity of the plant tissues, allowing high-resolution imaging that facilitated cell-specific localization of proanthocyanidins. A brown coloration was seen in proanthocyanidin-producing cells when plant tissues were embedded without DMACA staining and this was likely to have been due to non-enzymatic oxidation of proanthocyanidins and the formation of colored semiquinones and quinones.Conclusions
This paper presents a simple, high-resolution method for analysis of proanthocyanidin accumulation in organs, tissues and cells of two plant species with different patterns of proanthocyanidin accumulation, namely Lotus corniculatus (birdsfoot trefoil) and Trifolium repens (white clover). This technique was used to characterize cell type-specific patterns of proanthocyanidin accumulation in white clover flowers at different stages of development. 相似文献17.
Sandra Dèrozier Franck Samson Jean-Philippe Tamby Cécile Guichard Véronique Brunaud Philippe Grevet Séverine Gagnot Philippe Label Jean-Charles Leplé Alain Lecharny Sébastien Aubourg 《Plant methods》2011,7(1):1-10
Background
Efficient high throughput screening systems of useful mutants are prerequisite for study of plant functional genomics and lots of application fields. Advance in such screening tools, thanks to the development of analytic instruments. Direct analysis in real-time (DART)-mass spectrometry (MS) by ionization of complex materials at atmospheric pressure is a rapid, simple, high-resolution analytical technique. Here we describe a rapid, simple method for the genetic discrimination of intact Arabidopsis thaliana mutant seeds using metabolic profiling by DART-MS.Results
To determine whether this DART-MS combined by multivariate analysis can perform genetic discrimination based on global metabolic profiling, intact Arabidopsis thaliana mutant seeds were subjected to DART-MS without any sample preparation. Partial least squares-discriminant analysis (PLS-DA) of DART-MS spectral data from intact seeds classified 14 different lines of seeds into two distinct groups: Columbia (Col-0) and Landsberg erecta (Ler) ecotype backgrounds. A hierarchical dendrogram based on partial least squares-discriminant analysis (PLS-DA) subdivided the Col-0 ecotype into two groups: mutant lines harboring defects in the phenylpropanoid biosynthetic pathway and mutants without these defects. These results indicated that metabolic profiling with DART-MS could discriminate intact Arabidopsis seeds at least ecotype level and metabolic pathway level within same ecotype.Conclusion
The described DART-MS combined by multivariate analysis allows for rapid screening and metabolic characterization of lots of Arabidopsis mutant seeds without complex metabolic preparation steps. Moreover, potential novel metabolic markers can be detected and used to clarify the genetic relationship between Arabidopsis cultivars. Furthermore this technique can be applied to predict the novel gene function of metabolic mutants regardless of morphological phenotypes. 相似文献18.
Context
Objective identification of locations on transportation networks, where animal-vehicle collisions (AVC) occur more frequently than expected (hotspots), is an important step for the effective application of mitigation measures.Objectives
We introduce the KDE+ software which is a programmed version of the KDE+ method for effective identification of traffic accident hotspots. The software can be used in order to analyze animal-vehicle collision data.Methods
The KDE+ method is based on principles of Kernel Density Estimation (KDE). The symbol ‘+’ indicates that the method allows for the objective selection of significant clusters and for the ranking of the hotspots. It is also simultaneously applicable to an unlimited number of road segments.Results
We applied the KDE+ method to the entire Czech road network. The hotspots were ranked according to their significance. The resulting hotspots represent a short overall road length which should require a more detailed assessment in the field. The 100 most important clusters of AVC represent, for example, only 19.7 km of the entire road network (37,469 km).Conclusions
We present an objective method for hotspots identification which can be used for AVC data. This method is unique because it determines the significance level of hotspots in an objective way. The prioritization of hotspots allows a transportation manager to effectively allocate resources to a feasible number of identified hotspots. We describe the software, data preparation and present the KDE+ application to AVC data.20.
Hayley R. Tumas Brian M. Shamblin Mark Woodrey Nathan P. Nibbelink Richard Chandler Campbell Nairn 《Landscape Ecology》2018,33(9):1585-1601