Flow cytometry (FCM) is now the most widely used method to determine ploidy levels and genome size of plants. To get reliable estimates and allow reproducibility of measurements, the methodology should be standardized and follow the best practices in the field. In this article, we discuss instrument calibration and quality control and various instrument and acquisition settings (parameters, flow rate, number of events, scales, use of discriminators, peak positions). These settings must be decided before measurements because they determine the amount and quality of the data and thus influence all downstream analyses. We describe the two main approaches to raw data analysis (gating and histogram modeling), and we discuss their advantages and disadvantages. Finally, we provide a summary of best practice recommendations for data acquisition and raw data analysis in plant FCM.
A critical aspect for obtaining accurate, reliable, and high-resolution estimates of nuclear DNA content is the release of nuclei from the cytoplasm in sufficient amounts, while maintaining their integrity throughout the analysis, protecting their DNA from degradation by endonucleases, and enabling stoichiometric DNA staining. In embryophytes, the most common method consists of chopping the plant material with a sharp razor blade to release nuclei into an isolation buffer, filtering the homogenate, and staining the nuclei in buffered suspension with a fluorochrome of choice. Despite the recent description of alternative approaches to isolate nuclei, the chopping procedure remains the most widely adopted method, due to its simplicity, rapidity, and effectiveness. In this review article, we discuss the specifics of nuclei isolation buffers and the distorting effects that secondary metabolites may have in nuclear suspensions and how to test them. We also present alternatives to the chopping procedure, options for filtering and fluorochromes, and discuss the applications of these varied approaches. A summary of the best practices regarding the isolation of plant nuclei for the estimation of nuclear DNA content is also provided.
Pollen grains are the male gametophytes in a seed-plant life cycle. Their small, particulate nature and crucial role in plant reproduction have made them an attractive object of study using flow cytometry (FCM), with a wide range of applications existing in the literature. While methodological considerations for many of these overlap with those for other tissue types (e.g., general considerations for the measurement of nuclear DNA content), the relative complexity of pollen compared to single cells presents some unique challenges. We consider these here in the context of both the identification and isolation of pollen and its subunits, and the types of research applications. While the discussion here mostly concerns pollen, the general principles described here can be extended to apply to spores in ferns, lycophytes, and bryophytes. In addition to recommendations provided in more general studies, some recurring and notable issues related specifically to pollen and spores are highlighted.
- MeSH
- buněčné jádro MeSH
- ploidie MeSH
- průtoková cytometrie MeSH
- pyl * MeSH
- spory * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Local biodiversity trends over time are likely to be decoupled from global trends, as local processes may compensate or counteract global change. We analyze 161 long-term biological time series (15-91 years) collected across Europe, using a comprehensive dataset comprising ~6,200 marine, freshwater and terrestrial taxa. We test whether (i) local long-term biodiversity trends are consistent among biogeoregions, realms and taxonomic groups, and (ii) changes in biodiversity correlate with regional climate and local conditions. Our results reveal that local trends of abundance, richness and diversity differ among biogeoregions, realms and taxonomic groups, demonstrating that biodiversity changes at local scale are often complex and cannot be easily generalized. However, we find increases in richness and abundance with increasing temperature and naturalness as well as a clear spatial pattern in changes in community composition (i.e. temporal taxonomic turnover) in most biogeoregions of Northern and Eastern Europe.
- MeSH
- biodiverzita * MeSH
- ekosystém * MeSH
- klimatické změny MeSH
- Publikační typ
- časopisecké články MeSH
- metaanalýza MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Evropa MeSH
Polyploidisation has played an important role in plant diversification, and variation in ploidy level may be found not only between species of the same genus, but also within a single species. Although establishing the adaptive significance of polyploidy to explain the geographic distribution of cytotypes is challenging, the occurrence of different cytotypes in different ecological niches may suggest an adaptive role of genome duplication. We studied the adaptive significance of the geographic distribution of cytotypes across the entire distribution range of the endemic Erysimum mediohispanicum (Brassicaceae). For that, we have used climate variables, population elevation and soil properties to model ecological niches for the different cytotypes. In addition, we analysed the effect that ploidy level has on the floral phenotype. We found a clear geographic pattern in the distribution of cytotypes, with diploid individuals occurring in the southernmost part of the distribution range, while tetraploids were found in the northern area. A contact (mosaic) zone between both cytotypes was identified, but diploids and tetraploids occur in sympatry in only one population (although in a highly unbalanced proportion). Gene flow between different cytotypes seems to be negligible, as evident from an almost complete absence of triploids and other minority cytotypes. Niches occupied by both cytotypes showed subtle, but significant differences, even in the contact zone. Precipitation was higher in regions occupied by tetraploid individuals, which present wider corolla tubes and thinner but taller stalks than diploids. Our findings highlight the potential role of polyploidy in the ecological adaptation of E. mediohispanicum to both abiotic factors and biotic interactions.
BACKGROUND: Chickpea (Cicer arietinum L.) is a widely cropped pulse and an important source of proteins for humans. In Mediterranean regions it is predicted that drought will reduce soil moisture and become a major issue in agricultural practice. Nitrogen (N)-fixing bacteria and arbuscular mycorrhizal (AM) fungi have the potential to improve plant growth and drought tolerance. The aim of the study was to assess the effects of N-fixing bacteria and AM fungi on the growth, grain yield and protein content of chickpea under water deficit. RESULTS: Plants inoculated with Mesorhizobium mediterraneum or Rhizophagus irregularis without water deficit and inoculated with M. mediterraneum under moderate water deficit had significant increases in biomass. Inoculation with microbial symbionts brought no benefits to chickpea under severe water deficit. However, under moderate water deficit grain crude protein was increased by 13%, 17% and 22% in plants inoculated with M. mediterraneum, R. irregularis and M. mediterraneum + R. irregularis, respectively. CONCLUSION: Inoculation with N-fixing bacteria and AM fungi has the potential to benefit agricultural production of chickpea under water deficit conditions and to contribute to increased grain protein content. © 2017 Society of Chemical Industry.
- MeSH
- Bacteria růst a vývoj MeSH
- Cicer chemie růst a vývoj metabolismus mikrobiologie MeSH
- dusík metabolismus MeSH
- fyziologie bakterií * MeSH
- houby růst a vývoj fyziologie MeSH
- mykorhiza fyziologie MeSH
- očkovadla agrotechnická fyziologie MeSH
- rostlinné proteiny analýza metabolismus MeSH
- symbióza MeSH
- voda analýza metabolismus MeSH
- Publikační typ
- časopisecké články MeSH
