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Plant Reproduction Jun 2017Late-acting self-incompatibility (LSI) has been defined as a genetically controlled self-sterility mechanism that prevents seed set by selfing, despite normal pollen...
Late-acting self-incompatibility (LSI) has been defined as a genetically controlled self-sterility mechanism that prevents seed set by selfing, despite normal pollen tube growth and ovule penetration in self-pollinated pistils. In species of the Bignoniaceae with LSI, such as Handroanthus impetiginosus, the selfed pistils are characterized by a marked delay in ovule penetration, fertilization, and endosperm initiation, followed by uniform pistil abscission. This highlights the contentious possibility of a post-zygotic self-incompatibility system. However, previous studies were unable to confirm fusion of the sperm and egg cell nuclei in selfed ovules. In the present study, the cytology of the embryo sac, double fertilization, and pistil longevity was investigated in H. impetiginosus using comparative nuclei microspectrofluorometry of DAPI-stained sections of self- vs. unpollinated pistils. Differences in both pistil longevity and ovary size between self- and unpollinated flowers at the time of pistil abscission were significant. Zygotes with double the DNA content in their nuclei relative to unfertilized egg cell nuclei were verified in selfed ovules from the first day after pollination onward, and G1 karyogamy appeared to have occurred. Our cytological analysis clearly indicates that ovules of self-pollinated pistils in H. impetiginosus are fertilized before pistil abscission but no embryogenesis initiation occurs, which strongly supports the idea of a post-zygotic self-incompatibility mechanism.
Topics: Bignoniaceae; DNA, Plant; Flowers; Microspectrophotometry; Pollination; Self-Incompatibility in Flowering Plants
PubMed: 28303339
DOI: 10.1007/s00497-017-0300-7 -
Insects Aug 2023Quantifying the growth of entomopathogenic fungi is crucial for understanding their virulence and pathogenic potential. Traditional methods for determining growth, such...
Quantifying the growth of entomopathogenic fungi is crucial for understanding their virulence and pathogenic potential. Traditional methods for determining growth, such as biomass determination or colony growth area, are time-consuming and quantitatively and spatially limited in scope. In this study, we introduce a high-throughput method for rapidly measuring fungal growth using spectrophotometry in small-volume, liquid media cultures in 96-well microplates. Optical density (OD) changes were directly correlated with dry weight of samples for six isolates from three species of the genus to validate spectrophotometric growth measurements, and investigate species- and isolate-specific effects. We quantified fungal biomass from the microcultures by extracting, drying, and weighing mycelial mats. From the relationship established between OD and biomass, we generated standard curves for predicting biomass based on the OD values. The OD measurements clearly distinguished growth patterns among six isolates from three species. The logistic growth phase, as captured by the OD measurements, could be accurately assessed within a span of 80 h. Using isolates of , , and , this technique was demonstrated to be an effective, reproducible, and simple method for rapidly measuring filamentous fungal growth with high precision. This technique offers a valuable tool for studying the growth dynamics of entomopathogenic fungi and investigating the factors that influence their growth.
PubMed: 37623413
DOI: 10.3390/insects14080703 -
Journal of the American Chemical Society Jul 2023Single-molecule localization microscopy (SMLM) at cryogenic temperature opens new avenues to investigate intact biological samples at the nanoscale and perform...
Single-molecule localization microscopy (SMLM) at cryogenic temperature opens new avenues to investigate intact biological samples at the nanoscale and perform cryo-correlative studies. Genetically encoded fluorescent proteins (FPs) are markers of choice for cryo-SMLM, but their reduced conformational flexibility below the glass-transition temperature hampers efficient cryo-photoswitching. We investigated cryo-switching of rsEGFP2, one of the most efficient reversibly switchable fluorescent proteins at ambient temperature due to facile isomerization of the chromophore. UV-visible microspectrophotometry and X-ray crystallography revealed a completely different switching mechanism at ∼110 K. At this cryogenic temperature, on-off photoswitching involves the formation of two off-states in conformation with blue-shifted absorption relative to that of the protonated chromophore populated at ambient temperature. Only one of these off-states can be switched back to the fluorescent on-state by 405 nm light, while both of them are sensitive to UV light at 355 nm. Superior recovery to the fluorescent on-state by 355 nm light was confirmed at the single-molecule level. This suggests, as also shown by simulations, that employing 355 nm light in cryo-SMLM experiments using rsEGFP2 and possibly other FPs could improve the effective labeling efficiency achievable with this technique. The rsEGFP2 photoswitching mechanism discovered in this work adds to the panoply of known switching mechanisms in fluorescent proteins.
Topics: Temperature; Luminescent Proteins; Isomerism; Protein Conformation; Ultraviolet Rays
PubMed: 37389576
DOI: 10.1021/jacs.3c01500 -
Journal of AOAC International Sep 2018While color is arguably the most important optical property of evidential fibers, the actual dyestuffs responsible for its expression in them are, in forensic trace... (Review)
Review
While color is arguably the most important optical property of evidential fibers, the actual dyestuffs responsible for its expression in them are, in forensic trace evidence examinations, rarely analyzed and still less often identified. This is due, primarily, to the exceedingly small quantities of dye present in a single fiber as well as to the fact that dye identification is a challenging analytical problem, even when large quantities are available for analysis. Among the practical reasons for this are the wide range of dyestuffs available (and the even larger number of trade names), the low total concentration of dyes in the finished product, the limited amount of sample typically available for analysis in forensic cases, and the complexity of the dye mixtures that may exist within a single fiber. Literature on the topic of dye analysis is often limited to a specific method, subset of dyestuffs, or an approach that is not applicable given the constraints of a forensic analysis. Here, we present a generalized approach to dye identification that (1) combines several robust analytical methods, (2) is broadly applicable to a wide range of dye chemistries, application classes, and fiber types, and (3) can be scaled down to forensic casework-sized samples. The approach is based on the development of a reference collection of 300 commercially relevant textile dyes that have been characterized by a variety of microanalytical methods (HPTLC, Raman microspectroscopy, infrared microspectroscopy, UV-Vis spectroscopy, and visible microspectrophotometry). Although there is no single approach that is applicable to all dyes on every type of fiber, a combination of these analytical methods has been applied using a reproducible approach that permits the use of reference libraries to constrain the identity of and, in many cases, identify the dye (or dyes) present in a textile fiber sample.
Topics: Coloring Agents; Forensic Sciences; Humans; Microspectrophotometry; Reference Standards; Spectrophotometry; Spectrophotometry, Infrared; Spectrophotometry, Ultraviolet; Spectrum Analysis, Raman; Textiles
PubMed: 29669625
DOI: 10.5740/jaoacint.18-0052 -
Journal of Evolutionary Biology Jul 2015The dominant hypothesis for the evolutionary origin of snakes from 'lizards' (non-snake squamates) is that stem snakes acquired many snake features while passing through...
The dominant hypothesis for the evolutionary origin of snakes from 'lizards' (non-snake squamates) is that stem snakes acquired many snake features while passing through a profound burrowing (fossorial) phase. To investigate this, we examined the visual pigments and their encoding opsin genes in a range of squamate reptiles, focusing on fossorial lizards and snakes. We sequenced opsin transcripts isolated from retinal cDNA and used microspectrophotometry to measure directly the spectral absorbance of the photoreceptor visual pigments in a subset of samples. In snakes, but not lizards, dedicated fossoriality (as in Scolecophidia and the alethinophidian Anilius scytale) corresponds with loss of all visual opsins other than RH1 (λmax 490-497 nm); all other snakes (including less dedicated burrowers) also have functional sws1 and lws opsin genes. In contrast, the retinas of all lizards sampled, even highly fossorial amphisbaenians with reduced eyes, express functional lws, sws1, sws2 and rh1 genes, and most also express rh2 (i.e. they express all five of the visual opsin genes present in the ancestral vertebrate). Our evidence of visual pigment complements suggests that the visual system of stem snakes was partly reduced, with two (RH2 and SWS2) of the ancestral vertebrate visual pigments being eliminated, but that this did not extend to the extreme additional loss of SWS1 and LWS that subsequently occurred (probably independently) in highly fossorial extant scolecophidians and A. scytale. We therefore consider it unlikely that the ancestral snake was as fossorial as extant scolecophidians, whether or not the latter are para- or monophyletic.
Topics: Animals; Biological Evolution; Evolution, Molecular; Lizards; Molecular Sequence Data; Opsins; Phylogeny; Retina; Snakes
PubMed: 26012745
DOI: 10.1111/jeb.12663 -
Proceedings. Biological Sciences Sep 2016Previous investigations of vision and visual pigment evolution in aquatic predators have focused on fish and crustaceans, generally ignoring the cephalopods. Since the... (Comparative Study)
Comparative Study
Previous investigations of vision and visual pigment evolution in aquatic predators have focused on fish and crustaceans, generally ignoring the cephalopods. Since the first cephalopod opsin was sequenced in late 1980s, we now have data on over 50 cephalopod opsins, prompting this functional and phylogenetic examination. Much of this data does not specifically examine the visual pigment spectral absorbance position (λmax) relative to environment or lifestyle, and cephalopod opsin functional adaptation and visual ecology remain largely unknown. Here we introduce a new protocol for photoreceptor microspectrophotometry (MSP) that overcomes the difficulty of bleaching the bistable visual pigment and that reveals eight coastal coleoid cephalopods to be monochromatic with λmax varying from 484 to 505 nm. A combination of current MSP results, the λmax values previously characterized using cephalopod retinal extracts (467-500 nm) and the corresponding opsin phylogenetic tree were used for systematic comparisons with an end goal of examining the adaptations of coleoid visual pigments to different light environments. Spectral tuning shifts are described in response to different modes of life and light conditions. A new spectral tuning model suggests that nine amino acid substitution sites may determine the direction and the magnitude of spectral shifts.
Topics: Animals; Cephalopoda; Ecosystem; Phylogeny; Retinal Pigments; Rod Opsins
PubMed: 27629028
DOI: 10.1098/rspb.2016.1346 -
Small (Weinheim An Der Bergstrasse,... Feb 2024Thermochromic materials have been widely investigated due to their relevance in technological applications, including anti-counterfeiting materials, fashion accessories,...
Thermochromic materials have been widely investigated due to their relevance in technological applications, including anti-counterfeiting materials, fashion accessories, displays, and temperature sensors. While many organisms exhibit color changes, few studies have explored the potential of the responsive natural materials for temperature sensing, especially given the often limited and irreversible nature of these changes in live specimens. Here, it is shown that the hindwings of the blue-winged grasshopper Coloracris azureus can act as a reversible, power-free bio-thermometer, transitioning from blue to purple/red in a 30-100°C temperature range. Using microspectrophotometry, light microscopy and Raman microscopy, it is found that the blue color of the wings originates from pigmentary coloration, based on a complex of astaxanthin and proteins. The thermochromic shift from blue to red, induced by a temperature increase, is attributed to a denaturation of this carotenoprotein complex, upon which astaxanthin is released. This process is reversible upon a subsequent temperature decrease. The color changes are both swift and consistent upon temperature change, making the grasshopper's wings suitable as direct visual sensors on thermally dynamic, curved surfaces. The potential possibilities of sustainable, power-free temperature sensors or microthermometers based on biomaterials are demonstrated.
PubMed: 38366281
DOI: 10.1002/smll.202310193 -
Forensic Science International Mar 2019Edged weapons are frequently used to commit violent crimes (e.g., homicides, assaults or sequestrations). Following a stabbing, a fibre transfer might occur from the...
Edged weapons are frequently used to commit violent crimes (e.g., homicides, assaults or sequestrations). Following a stabbing, a fibre transfer might occur from the damaged fabric to the blade. It is crucial to investigate whether the target fibres were transferred to the blade as a result of the stabbing and if they are in association with the offence or whether they originate from another unrelated event (i.e., part of the background). The aim of this study is to explore the background population of fibres recovered on knife blades (police seizures and everyday knives) to help the interpretation of the results stemming from the analysis of the fibre evidence in crime cases involving the use of a bladed weapon. A population study was carried out to learn about the variety, the distribution and the size of the fibre population present on knife blades. After recovery using tapes lifting, stereoscopic examinations were performed to classify around 1500 fibres according to their colour and generic type. The most common colour/type combinations were: colourless/white cotton (31%), blue cotton (16%) and grey cotton fibres (11%). As blue fibres represent one of the most common colour group, they were selected to carry out a colour block study (based on microscopy observations and microspectrophotometry UV-vis spectra) to investigate the number and the size of the groups of indistinguishable fibres present on each blade. Only 30% of the knife blades presented more than one blue cotton fibre. The number of groups found per knife varied between 1 and 3, each group containing between 2 and 6 fibres. Findings revealed an important difference between the background fibre populations in terms of quantity of fibres, their morphology and spatial distributions - and the population of fibres transferred in a stabbing. The collected data can be used in a Bayesian approach to help with the interpretation of results.
PubMed: 30731332
DOI: 10.1016/j.forsciint.2019.01.006 -
Forensic Science International Jan 2021Soil is a very important type of trace evidence. The iron content of soil is of great significance in distinguishing soil types, discriminating among different soils,...
Soil is a very important type of trace evidence. The iron content of soil is of great significance in distinguishing soil types, discriminating among different soils, and tracing soils. However, conventional methods for analyzing the iron content of soil are expensive, laborious, and time-consuming. Previous studies have shown that the color of soil correlates well with its hematite content. This article thus deals with the indirect determination of iron content using soil color as a proxy. Soil color measurements were conducted using microspectrophotometry (MSP), and resulting data were transformed into chromaticity value (L*, a*, and b*). Predictions using the redness index in conjunction with a linear regression model were compared with those using the chromaticity value and a back propagation neural network (BPNN) model. The influences of different modeling conditions on the modeling accuracy were compared, and more accurate predictions were achieved when the iron content was higher than 2.13%. The BPNN model produced predictions with R and RMSE values of 0.955 and 0.336%, which were better than the predictions of the linear regression model (R: 0.859, RMSE, 1.07%). We thus demonstrated that MSP can be used for fast, accurate, and non-destructive measurements of soil color and prediction of its iron content. Although the results may not be as precise as conventional laboratory analysis, they still provide more information with acceptable accuracy, which should have promising applications in forensic applications.
PubMed: 33278697
DOI: 10.1016/j.forsciint.2020.110600 -
Protein and Peptide Letters 2016This review describes the use of single crystal UV-Visible Absorption micro-Spectrophotometry (UV-Vis AS) to enhance the design and execution of X-ray crystallography... (Review)
Review
This review describes the use of single crystal UV-Visible Absorption micro-Spectrophotometry (UV-Vis AS) to enhance the design and execution of X-ray crystallography experiments for structural investigations of reaction intermediates of redox active and photosensitive proteins. Considerations for UV-Vis AS measurements at the synchrotron and associated instrumentation are described. UV-Vis AS is useful to verify the intermediate state of an enzyme and to monitor the progression of reactions within crystals. Radiation induced redox changes within protein crystals may be monitored to devise effective diffraction data collection strategies. An overview of the specific effects of radiation damage on macromolecular crystals is presented along with data collection strategies that minimize these effects by combining data from multiple crystals used at the synchrotron and with the X-ray free electron laser.
Topics: Crystallography, X-Ray; Macromolecular Substances; Models, Molecular; Spectrophotometry, Ultraviolet; Synchrotrons; X-Rays
PubMed: 26740326
DOI: 10.2174/0929866523666160107115015