Individual Pollen Research Articles

Understanding the Systematic Implications of Pollinium Morphology in the Aeridinae Subtribe (Orchidaceae)

Abstract— Molecular phylogenetic studies have been the basis for major clades proposed for the Aeridinae subtribe of the Orchidaceae family. However, systematic morphological characterization has yet to be conducted. We carried out this study on Aeridinae pollinium morphologies with an emphasis on aperture type and pollinium sculpturing to determine the systematic significance of these characteristics in the context of molecular phylogenetics. The results indicated that most Aeridinae species have cleft type pollinium apertures and that few have porate type or no aperture. The shapes of most of the pollinia were flat but this was not consistent at the genus level. Pollinium length ranged from 204 to 2606 µm, and the width from 178 to 1524 µm. Long and wide pollinia were commonly found in primitive groups, while short and narrow pollinia were commonly found in evolutionarily derived groups. The shapes and sizes of the individual units of the pollinia varied from uniform to variable, with a few exceptions. The individual units of most the pollinia were tetrads, and individual pollen grains were easily distinguished within the tetrad. The degree to which individual pollen grains could be differentiated is constant at the clade level except for Aerides rosea. The exine sculpturing characteristics were consistent in each clade except in the Cleisostoma and Phalaenopsis clades. The pollinium sculpturing has a somewhat clear pattern of change that was consistent with the molecular phylogeny, with a few exceptions. In conclusion, these pollinium characteristics themselves are not systematic, but they can serve as useful characteristics for determining the major systematic groups in the subtribe Aeridinae.

Trees Are a Major Foraging Resource for Honeybees in the City.

Large cities are typically characterized by a mosaic of green spaces that hold a remarkable variety of native and "exotic" plants. Urban beekeeping has gained increasing popularity. In order to characterize the "urban" in the honey, pollen diversity in 50 honey samples from 18 apiary locations in Vienna, Austria, was microscopically analyzed. The relative abundances of each plant taxon were determined by counting out 500 individual pollen grains per sample. In total, 202 taxa could be identified, with a median of 46 per sample. Taxa richness and diversity differed significantly across three years but did not so between urban and suburban apiaries. Despite trees comprising only roughly a quarter of all taxa, the amount of tree pollen was disproportionally high. The invasive Ailanthus altissima was predominant in 15 out of 50 samples. Other important non-native and/or ornamental trees included Sophora japonica, Gleditsia triacanthos, Castanea sativa, Koelreuteria paniculata, and Liriodendron tulipifera. Urban honey from Central Europe may typically comprise pollen taxa from Europe, East Asia, and North America alike. The results of this study show that intentionally planted, managed urban green spaces can support stable foraging resources for pollinators in cities.

MIKC*-type MADS transcription factors control JINGUBANG expression and the degree of pollen dormancy in Arabidopsis.

While pollen dormancy has been proposed to play a necessary role in sexual reproduction, it remains poorly understood. Here, we used traditional pollen germination assays to characterize dormancy. Our results underscore variation in the degree of dormancy between individual pollen grains. In addition, we provide evidence that JINGUBANG (JGB), previously defined as a negative regulator of pollen germination in Arabidopsis (Arabidopsis thaliana), is responsible for the uneven degrees of pollen dormancy, as asynchronous pollen germination in vitro reflected varied expression levels of JGB. We identified five cis-acting elements, including four CArG-boxes and the previously uncharacterized element ERE7, as essential for the initiation and enhancement of JGB expression. A 10-bp sequence between CArG-box 3 and ERE7, likely the result of an inverse DNA loop formed between CArG-box 3 and CArG-box 4, was required for robust gene expression. In addition, the pollen-specific AtMIKC*-type MADS transcription factors AGAMOUS-LIKE 30 (AGL30), AGL65, AGL66, AGL94, and AGL104 activated JGB transcription. Notably, the transactivation levels differed among the obligate AtMIKC* heterodimers tested. Our results indicate that distinct AtMIKC* complexes formed in individual pollen grains direct pollen dormancy to uneven degrees, which is likely an adaptive trait that ensures broader pollen dispersal under adverse environmental conditions.

Pollinator-mediated selection on Krameria oil flowers: a flower-pollinator fit adaptation to an atypical oil-collecting behaviour?

Spatial variation in plant-pollinator interactions is a key driver of floral trait diversification. A so far overlooked qualitative aspect of this variation is the behavioural component on flowers that relates to the pollinator fit. We tested the hypothesis that variation in pollinator behaviour influences the geographical pattern of phenotypic selection across the distribution range of the oil-producing Krameria grandiflora (Krameriaceae). This variation mainly involves the presence or absence of flag petal grasping, which is only performed by representatives of Centris (Centridini, Apidae), an oil-collecting bee group highly associated with Krameriaceae pollination. We quantified variation in floral traits and fitness and estimated pollinator-mediated selection in five populations at a large geographical scale comprising the entire species range. In each population, we sampled individual pollen arrival and germination as a fitness measure, indicating pollination success and pollination performance, which was then relativized and regressed on standardized flower-pollinator fit (flag-stigma distance), advertisement (sepal length) and reward (oil volume) traits. This generated mean-scaled selection gradients used to calculate geographical selection dispersion. Unexpectedly, stronger selection was detected on the flower-pollinator fit trait in populations highly associated with the absence of flag petal grasping. Geographical variation in selection was mainly attributed to differential selection on the flag-stigma distance generating a selection mosaic. This may involve influences of a spatial variation in pollinator behaviour as well as composition and morphology. Our results show the adaptive significance of the specialized flag petals of Krameria in the absence of the grasping behaviour and highlight the contribution of geographical variation in pollinator behaviour on flowers in driving selection mosaics, with implications for floral evolution, adaptation to pollinator fit and phenotypic diversity in specialized systems.

Greater difference between airborne and flower pollen chemistry, than between pollen collected across a pollution gradient in the Netherlands

The prevalence in allergic diseases has increased considerably in the past decades. An important trigger of the symptoms of allergic rhinitis (hay fever) is the pollen of wind-pollinating plants. This pollen is developed by plants and is released into the air where it gets exposed to environmental influences and air pollution. We investigated the chemical changes to pollen that occur after release from the flower in a rural (Veluwe) and an urban (Amsterdam) site in the Netherlands using Fourier Transform Infrared (FTIR) spectroscopy. During the spring/summer of 2020 (during the COVID pandemic) the pollen of nine taxa (Alnus, Betula, Fagus, Fraxinus, Pinus, Plantago, Poaceae, Quercus and Salix) were collected directly from flowers and the air (using a mobile sampler). FTIR spectra were obtained for multiple individual pollen grains for each taxa. The spectra obtained from airborne pollen collected at the rural vs. urban sites did not show any statistical difference. This is possibly a result of a reduced difference in pollutant concentrations between the two sites due to the COVID-19-lockdown measures were in place. However, consistent differences in the FTIR spectra recovered from airborne vs. flower pollen were recorded for all pollen taxa. After the release from the flower the chemical composition of the pollen changed: (i) polysaccharides are converted to monosaccharides; (ii) protein concentration and/or nitration/oxidation level is altered; (iii) lipids are modified and/or reduced in concentration. These changes may alter the allergenicity of the pollen and suggest that further work on the allergenic nature of airborne pollen is required.

FTIR spectra from grass pollen: A quest for species-level resolution of Poaceae and Cerealia-type pollen grains

Palynological analysis based on spore and pollen morphology is well established in the field of palaeo-environmental reconstruction but is currently not fully exploited for understanding the history and development of cereal cultivation due to difficulties in visually differentiating between grass species (Poaceae). Here we employ a chemotaxonomic approach, by examining the chemical differences among Poaceae taxa, based on Fourier-transform infrared (FTIR) microspectroscopy data to overcome problems associated with morphological similarities across the Poaceae family. FTIR spectra of untreated and acetolysed pollen from 19 Poaceae taxa were used in our study. We used both populations and individual pollen grains to explore how we can minimise the effect of Mie scattering (spectral distortions caused by scattering of the incident IR beam) on spectra from individuals. Random forest classification algorithms were applied to explore our ability to differentiate taxa at the species level. We found that pollen grains treated with acetolysis yield better classification results (86% for individuals and 97% for populations) compared to untreated samples (65.7% for individuals and 83% for populations), since they are less affected by Mie scattering. The high classification success at species level on acetolysed individual pollen grains suggests that our chemotaxonomic method holds substantial promise in numerous areas of grass and in particular cereal pollen research, including elucidating the history of agriculture.

Pollen-based reconstructions of Holocene climate trends in the eastern Mediterranean region

Abstract. There has been considerable debate about the degree to which climate has driven societal changes in the eastern Mediterranean region, partly through reliance on a limited number of qualitative records of climate changes and partly reflecting the need to disentangle the joint impact of changes in different aspects of climate. Here, we use tolerance-weighted, weighted-averaging partial least squares to derive reconstructions of the mean temperature of the coldest month (MTCO), mean temperature of the warmest month (MTWA), growing degree days above a threshold of 0 ∘C (GDD0), and plant-available moisture, which is represented by the ratio of modelled actual to equilibrium evapotranspiration (α) and corrected for past CO2 changes. This is done for 71 individual pollen records from the eastern Mediterranean region covering part or all of the interval from 12.3 ka to the present. We use these reconstructions to create regional composites that illustrate the long-term trends in each variable. We compare these composites with transient climate model simulations to explore potential causes of the observed trends. We show that the glacial–Holocene transition and the early part of the Holocene was characterised by conditions colder than the present. Rapid increases in temperature occurred between ca. 10.3 and 9.3 ka, considerably after the end of the Younger Dryas. Although the time series are characterised by centennial to millennial oscillations, the MTCO showed a gradual increase from 9 ka to the present, consistent with the expectation that winter temperatures were forced by orbitally induced increases in insolation during the Holocene. The MTWA also showed an increasing trend from 9 ka and reached a maximum of ca. 1.5 ∘C greater than the present at ca. 4.5 and 5 ka, followed by a gradual decline towards present-day conditions. A delayed response to summer insolation changes is likely a reflection of the persistence of the Laurentide and Fennoscandian ice sheets; subsequent summer cooling is consistent with the expected response to insolation changes. Plant-available moisture increased rapidly after 11 ka, and conditions were wetter than today between 10 and 6 ka, but thereafter, α declined gradually. These trends likely reflect changes in atmospheric circulation and moisture advection into the region and were probably too small to influence summer temperature through land–surface feedbacks. Differences in the simulated trajectory of α in different models highlight the difficulties in reproducing circulation-driven moisture advection into the eastern Mediterranean.

Six-Food Elimination Diet Is Less Effective During Pollen Season in Adults With Eosinophilic Esophagitis Sensitized to Pollens.

The role of inhaled and swallowed aeroallergens in treatment outcomes of adult patients with eosinophilic esophagitis (EoE) is unclear. We hypothesized that the pollen season contributes to the failure of the 6-food elimination diet (SFED) in EoE. We compared outcomes of patients with EoE who underwent SFED during vs outside of the pollen season. Consecutive adult patients with EoE who underwent SFED and skin prick test (SPT) for birch and grass pollen were included. Individual pollen sensitization and pollen count data were analyzed to define whether each patient had been assessed during or outside of the pollen season after SFED. All patients had active EoE (≥15 eosinophils/high-power field) before SFED and adhered to the diet under the supervision of a dietitian. Fifty-eight patients were included, 62.0% had positive SPT for birch and/or grass, whereas 37.9% had negative SPT. Overall, SFED response was 56.9% (95% confidence interval, 44.1%-68.8%). When stratifying response according to whether the assessment had been performed during or outside of the pollen season, patients sensitized to pollens showed significantly lower response to SFED during compared with outside of the pollen season (21.4% vs 77.3%; P = 0.003). In addition, during the pollen season, patients with pollen sensitization had significantly lower response to SFED compared with those without sensitization (21.4% vs 77.8%; P = 0.01). Pollens may have a role in sustaining esophageal eosinophilia in sensitized adults with EoE despite avoidance of trigger foods. The SPT for pollens may identify patients less likely to respond to the diet during the pollen season.

Pollen and asthma morbidity in Atlanta: A 26-year time-series study

BackgroundCompared to many environmental risk factors, the relationship between pollen and asthma is understudied, including how associations may differ by pollen type and between subgroups, and how associations may be changing over time. ObjectivesWe evaluated the association between ambient pollen concentrations and emergency department (ED) visits for asthma and wheeze in Atlanta, Georgia during 1993–2018. We estimated overall associations for 13 individual pollen taxa, as well as associations by decade, race, age (5–17, 18–64, 65+), and insurance status (Medicaid vs non-Medicaid). MethodsSpeciated pollen data were acquired from Atlanta Allergy & Asthma, a nationally certified pollen counting station. ED visit data were obtained from individual hospitals and from the Georgia Hospital Association. We performed time-series analyses using quasi-Poisson distributed lag models, with primary analyses assessing 3-day (lag 0–2 days) pollen levels. Models controlled for day of week, holidays, air temperature, month, year, and month-by-year interactions. ResultsFrom 1993 to 2018, there were 686,259 ED visits for asthma and wheeze in the dataset, and the number of ED visits increased over time. We observed positive associations of asthma and wheeze ED visits with nine of the 13 pollen taxa: trees (maple, birch, pine, oak, willow, sycamore, and mulberry), two weeds (nettle and pigweed), and grasses. Rate ratios indicated 1–8% increases in asthma and wheeze ED visits per standard deviation increases in pollen. In general, we observed stronger associations in the earliest period (1993–2000), in younger people, and in Black patients; however, results varied by pollen taxa. ConclusionsSome, but not all, types of pollen are associated with increased ED visits for asthma/wheeze. Associations are generally higher in Black and younger patients and appear to have decreased over time.

Single cell ICP-MS as a powerful analytical tool to determine metal content in individual pollen grains

SC ICP MS analysis of Fe and Mn in pollen samples.

Towards European automatic bioaerosol monitoring: Comparison of 9 automatic pollen observational instruments with classic Hirst-type traps

To benefit allergy patients and the medical practitioners, pollen information should be available in both a reliable and timely manner; the latter is only recently possible due to automatic monitoring. To evaluate the performance of all currently available automatic instruments, an international intercomparison campaign was jointly organised by the EUMETNET AutoPollen Programme and the ADOPT COST Action in Munich, Germany (March–July 2021).The automatic systems (hardware plus identification algorithms) were compared with manual Hirst-type traps. Measurements were aggregated into 3-hourly or daily values to allow comparison across all devices. We report results for total pollen as well as for Betula, Fraxinus, Poaceae, and Quercus, for all instruments that provided these data. The results for daily averages compared better with Hirst observations than the 3-hourly values. For total pollen, there was a considerable spread among systems, with some reaching R2 > 0.6 (3 h) and R2 > 0.75 (daily) compared with Hirst-type traps, whilst other systems were not suitable to sample total pollen efficiently (R2 < 0.3). For individual pollen types, results similar to the Hirst were frequently shown by a small group of systems. For Betula, almost all systems performed well (R2 > 0.75 for 9 systems for 3-hourly data). Results for Fraxinus and Quercus were not as good for most systems, while for Poaceae (with some exceptions), the performance was weakest. For all pollen types and for most measurement systems, false positive classifications were observed outside of the main pollen season. Different algorithms applied to the same device also showed different results, highlighting the importance of this aspect of the measurement system. Overall, given the 30 % error on daily concentrations that is currently accepted for Hirst-type traps, several automatic systems are currently capable of being used operationally to provide real-time observations at high temporal resolutions. They provide distinct advantages compared to the manual Hirst-type measurements.

Pollen collection by honey bee hives in almond orchards indicate diverse diets

Almond is one of the world's most economically valuable crops and many varieties require cross pollination for optimal fruit set. For this reason, western honey bee (Apis mellifera L.) hives are often placed in almond orchards. However, little is known about the usage of almond and other pollen sources by individual hives during almond bloom. Here, we investigated the timing, identity and quantity of pollen collection associated with almond floral abundance and spatial location of individual hives by sampling 440 individual pollen tray samples and counting 45,072 pollen grains from 13,200 pollen pellets collected from 80 individual hives across the flowering season in Victoria, south-eastern Australia. A large proportion of hives collected non-almond pollen in addition to almond pollen (63/80 = 79%). The weight of almond pollen collected by the hives at each sampling time was positively related to the number of concurrently open almond flowers. However, non-almond pollen richness and abundance was not related to the number of almond flowers but had a positive relationship with the weight of almond pollen collected. There was no relationship between the distance among hives and identity of pollen collected. Yet, three plant families in the study area were found to account for a high percentage of the non-almond pollen collected, Euphorbiaceae, Fabaceae and Asteraceae. Understanding crop and non-crop pollen collection could inform honey bee diet needs and identify the plant species of importance to inform best practice bee management during almond flowering.

Pollen interference emerges as a property from agent-based modelling of pollen competition in Arabidopsis thaliana

Abstract Differences in pollen performance, often revealed during pollen competition, have long been recognized as evolutionarily significant and agriculturally important. Though we have sophisticated models for the growth of individual pollen tubes, we have few robust models for larger-scale pollen competition, a process that has been linked with inbreeding avoidance, sexual selection, reproductive barrier reinforcement and speciation. Here we use existing data on pollen performance traits to develop an agent-based model of pollen competition. We calibrate our model parameters to empirical data found in the literature of seed siring proportions from mixed pollinations and pollen tube length distributions from single-accession pollinations. In this model, parameters that influence pollen tube movement and sensing of ovules were found to be primary factors in competition. Our model also demonstrates that interference competition emerges as a property of pollen competition, and suggests a potential mechanism for this phenomenon. This study integrates pollen performance measures with mathematical modelling conducted on a simplified and accessible system. This represents the first mechanistic agent-based model for pollen competition. Our model may be extended to predict seed siring proportions for other accessions of Arabidopsis thaliana given data on their pollen performance traits.

The potential of multispectral imaging flow cytometry for environmental monitoring.

Environmental monitoring involves the quantification of microscopic cells and particles such as algae, plant cells, pollen, or fungal spores. Traditional methods using conventional microscopy require expert knowledge, are time-intensive and not well-suited for automated high throughput. Multispectral imaging flow cytometry (MIFC) allows measurement of up to 5000 particles per second from a fluid suspension and can simultaneously capture up to 12 images of every single particle for brightfield and different spectral ranges, with up to 60x magnification. The high throughput of MIFC has high potential for increasing the amount and accuracy of environmental monitoring, such as for plant-pollinator interactions, fossil samples, air, water or food quality that currently rely on manual microscopic methods. Automated recognition of particles and cells is also possible, when MIFC is combined with deep-learning computational techniques. Furthermore, various fluorescence dyes can be used to stain specific parts of the cell to highlight physiological and chemical features including: vitality of pollen or algae, allergen content of individual pollen, surface chemical composition (carbohydrate coating) of cells, DNA- or enzyme-activity staining. Here, we outline the great potential for MIFC in environmental research for a variety of research fields and focal organisms. In addition, we provide best practice recommendations.

MINIMISATION OF RISKS ASSOCIATED WITH THE USE OF POLLEN-BASED MEDICINES, AT THE STAGE OF POLLEN COLLECTION

The aim of the study is the elaboration of Rules for Harvesting/Collecting of Pollen to minimize the risks associated with the use of pollen-based medicinal products.Materials and methods. The following electronic resources were used in the study: PubMed, Medline, ScienceDirect, Web of Science, Scopus, Google Scholar, eLibrary, World Allergy Organization, Cochrane Database, Stallergenesgreer, Allergenscienceandconsulting, Pharmacopoeia, Fda.gov, fs.usda.gov, Ema.europa.eu. The analysis covered the period from January 1, 2010 until December 31, 2021.Results. Currently, there are some general requirements for the quality of pollen in Russia, but there are no controls or standardised procedures for harvesting, drying, and purification of pollen. The USA and EU also lack established qualification programmes for pollen-collecting companies and/or individual pollen collectors. Regulatory authorities establish requirements only for visual control of raw materials or delegate responsibility to the manufacturer. The analysis of the existing regulatory documentation revealed lack of requirements for collection, storage, and processing of pollen used as the raw material for the production of allergen products. This calls for the elaboration of appropriate regulatory documents. The authors have compiled the Rules for Harvesting/Collection of Pollen, which include 6 parts. The Rules are intended for individuals directly involved in harvesting/collection of pollen, and contain requirements for pollen collectors, the process of pollen collection, documentation, storage, and transportation.Conclusion. The authors have prepared the Rules for Harvesting/Collecting of Pollen, which include 6 parts. The Rules cover the whole process of pollen collection and all related processes. The implementation of this document will improve the process of pollen collection, thus reducing the risks associated with the use of pollen-based medicines. Further studies will assess the impact of the pollen quality on the safety of medicinal products.

Why should we care about high temporal resolution monitoring of bioaerosols in ambient air?

This is the first time that atmospheric concentrations of individual pollen types have been recorded by an automatic sampler with 1-hour and sub-hourly resolution (i.e. 1-minute and 1-second data). The data were collected by traditional Hirst type methods and state-of the art Rapid-E real-time bioaerosol detector. Airborne pollen data from 7 taxa, i.e. Acer negundo, Ambrosia, Broussonetia papyrifera, Cupressales (Taxaceae and Cupressaceae families), Platanus, Salix and Ulmus, were collected during the 2019 pollen season in Novi Sad, Serbia. Pollen data with daily, hourly and sub-hourly temporal resolution were analysed in terms of their temporal variability. The impact of turbulence kinetic energy (TKE) on pollen cloud homogeneity was investigated. Variations in Seasonal Pollen Integrals produced by Hirst and Rapid-E show that scaling factors are required to make data comparable. Daily average and hourly measurements recorded by the Rapid-E and Hirst were highly correlated and so examining Rapid-E measurements with sub-hourly resolution is assumed meaningful from the perspective of identification accuracy. Sub-hourly data provided an insight into the heterogenous nature of pollen in the air, with distinct peaks lasting ~5–10 min, and mostly single pollen grains recorded per second. Short term variations in 1-minute pollen concentrations could not be wholly explained by TKE. The new generation of automatic devices has the potential to increase our understanding of the distribution of bioaerosols in the air, provide insights into biological processes such as pollen release and dispersal mechanisms, and have the potential for us to conduct investigations into dose-response relationships and personal exposure to aeroallergens.

Use of pollen mixes for skin prick testing is efficient but misses sensitization to some included pollens

At Scripps Clinic pollen mixes are routinely used in a skin prick screening panel to assess for evidence of sensitization to aeroallergens in patients with suspect symptoms. In those unlikely to pursue aeroallergen immunotherapy identification of sensitization to individual pollens may be unnecessary. Use of mixes is time and cost saving. Notably the concentration of each individual pollen is less in these mixes with dilution based on the total number of pollens included. As a quality measure we seek to assess whether use of pollen mixes yield concordant results when compared to individual pollens testing.

757Using smartphone technology to characterise associations between respiratory symptoms and pollen

Abstract Background Pollen is a well-established trigger of asthma and allergic rhinoconjunctivitis, yet key gaps in our understanding remain. These include knowledge of concentration thresholds for symptoms, exposure-response associations through time, and the potential for interactions with other environmental stressors such as air pollution. Smartphone technology offers an opportunity to address these challenges using large datasets that capture individual symptoms in real time. Methods We analysed 44,820 symptom reports logged by 2,272 users of the AirRater app over four years to evaluate associations between daily respiratory symptoms and atmospheric concentrations of pollen in Tasmania, Australia. We used case time series, a novel methodology developed for app-sourced data. We adjusted for seasonality and meteorology and tested for interactions with particulate pollution (PM2.5). Results There was a non-linear association between pollen concentrations and respiratory symptoms for up to three days following exposure. Risk ratios (RR) were greatest on the same day, for total pollen increased steeply to a RR of 1.31 (95% CI: 1.26-1.37) at a concentration of 50 grains/m3 before plateauing. Associations with individual pollen taxa showed similar non-linear trends. There was an interaction with PM2.5, with effect estimates significantly higher when PM2.5 was &amp;gt;50 µg/m3 (p for interaction &amp;lt; 0.001). Conclusions The association between respiratory symptoms and airborne pollen was non-linear, greatest in magnitude on the day of exposure, and synergistic with air pollution. Key messages Smartphone symptom tracking offers a useful means of assessing dose-response relationships in environmental epidemiology.

Biomechanics of pollen pellet removal by the honey bee

Honey bees (Apis mellifera) carry pollen back to their hive by mixing it with nectar and forming it into a pellet. The pellet must be firmly attached to their legs during flight, but also easily removable when deposited in the hive. How does the honey bee achieve these contrary aims? In this experimental study, we film honey bees removing pollen pellets and find they peel them off at speeds 2-10 times slower than their typical grooming speeds. Using a self-built pollen scraper, we find that slow removal speeds reduce the force and work required to remove the pellet under shear stress. Creep tests on individual pollen pellets revealed that pollen pellets are viscoelastic materials characterized by a Maxwell model with long relaxation times. The relaxation time enables the pellet to remain a solid during both transport and removal. We hope that this work inspires further research into viscoelastic materials in nature.

High-throughput measuring of meiotic recombination rates in barley pollen nuclei using Crystal Digital PCRTM.

Breeding exploits novel allelic combinations assured by meiotic recombination. Barley (Hordeum vulgare) single pollen nucleus genotyping enables measurement of meiotic recombination rates in gametes before fertilization without the need for segregating populations. However, so far, established methods rely on whole-genome amplification of every single pollen nucleus due to their limited DNA content, thus restricting the number of analyzed samples. In this study, high-throughput measurements of meiotic recombination rates in barley pollen nuclei without whole-genome amplification were performed through a Crystal Digital PCRTM -based genotyping assay. Meiotic recombination rates within two centromeric and two distal chromosomal intervals were measured in hybrid plants by genotyping a total of >42000 individual pollen nuclei (up to 4900 nuclei analyzed per plant). Determined recombination frequencies in pollen nuclei were similar to frequencies in segregating populations. We improved the efficiency of the genotyping by pretreating the pollen nuclei with a thermostable restriction enzyme. Additional opportunities for a higher sample throughput and a further increase of the genotyping efficiency are presented and discussed. Taken together, single barley pollen nucleus genotyping based on Crystal Digital PCRTM enables reliable, rapid and high-throughput meiotic recombination measurements within defined chromosomal intervals of intraspecific hybrid plants. The successful encapsulation of nuclei from a range of species with different nuclear and genome sizes suggests that the proposed method is broadly applicable to genotyping single nuclei.