Global warming has already seen a radical change in temperature regimes in Bangladesh. This review provides the first up-to-date perspective and detailed analysis of wheat research in Bangladesh and the impact that global warming will have on its agriculture, especially wheat farming.
Background and aims
The most fundamental activity of the people of Bangladesh is agriculture. Modelling projections for Bangladesh indicate that warmer temperatures linked to climate change will severely reduce the growth of various winter crops (wheat, boro rice, potato and winter vegetables) in the north and central parts. In summer, crops in south-eastern parts of the country are at risk from increased flooding as sea levels increase.
Wheat is one of the most important winter crops and is temperature sensitive and the second most important grain crop after rice. In this review, we provide an up-to-date and detailed account of wheat research of Bangladesh and the impact that global warming may have on agriculture, especially wheat production. Although flooding is not of major importance or consequence to the wheat crop at present, some perspectives are provided on this stress since wheat is flood sensitive and the incidence of flooding is likely to increase.
This information and projections will allow wheat breeders to devise new breeding programmes to attempt to mitigate future global warming. We discuss what this implies for food security in the broader context of South Asia.
Cassava flowering with emphasis on flowering pattern, morphology and phenology; pollen biology on viability and dimorphism, and histology on male and female gametophyte development are demonstrated. Reduced pollen viability at anthesis and the existence of pollen tri-morphism are the key findings.
Background and aims
Cassava (Manihot esculenta), a major food staple in the tropics and subtropics, thrives even in environments undergoing threatening climate change. To satisfy the increasing demand for crop improvement and overcome the limitations of conventional breeding, the introduction of inbreeding techniques such as the production of doubled haploid lines via androgenesis or gynogenesis offers advantages. However, comprehensive studies on cassava flower bud biology or structural development are lacking and precise structural and biological information is a prerequisite to enhance the efficiency of these techniques.
The floral biology of three selected cassava lines was studied, focusing on morphology, phenology and pollen biology (quantity, viability and dimorphism). Histological studies were also conducted on microsporogenesis/microgametogenesis and megasporogenesis/megagameto-genesis to generate precise developmental data for these lines.
Male and female cyathia have distinct developmental phases. Pollen viability was high during immature stages of plant development; however, pollen mortality was common at later stages. Pollen trimorphism in male gametophytes towards the larger or smaller pollen size, as compared with normal size, was observed. Ten characteristic events were identified in male gametogenesis and six in female gametogenesis that were correlated with flower bud diameter. Male gametophyte diameter at different developmental stages was also determined.
Results indicate that the three lines did not differ significantly, except regarding a few morphological aspects such as plant height, flower colour and number of male cyathia. Pollen grains were initially viable, but viability decreased drastically at later stages of growth. Abnormal meiosis or mitosis triggered pollen trimorphism. The demonstrated sequential events of reproductive development generated valuable information at the cellular level, which will help close the current information gap for cassava improvement via breeding programmes and doubled haploid plant production.
Free auxin and expression of auxin-related genes were measured at ripening in MADS8 suppressed apple fruit. It was found that the delayed ripening in these fruit was associated with high auxin and changes in auxin homeostasis and response genes.
Background and aims
Fruit ripening is an important developmental trait in fleshy fruits, making the fruit palatable for seed dispersers. In some fruit species, there is a strong association between auxin concentrations and fruit ripening. We investigated the relationship between auxin concentrations and the onset of ethylene-related ripening in Malus × domestica (apples) at both the hormone and transcriptome levels.
Transgenic apples suppressed for the SEPALLATA1/2 (SEP1/2) class of gene (MADS8/9) that showed severely reduced ripening were compared with untransformed control apples. In each apple type, free indole-3-acetic acid (IAA) concentrations were measured during early ripening. The changes observed in auxin were assessed in light of global changes in gene expression.
It was found that mature MADS8/9-suppressed apples had a higher concentration of free IAA. This was associated with increased expression of the auxin biosynthetic genes in the indole-3-acetamide pathway. Additionally, in the MADS8/9-suppressed apples, there was less expression of the GH3 auxin-conjugating enzymes. A number of genes involved in the auxin-regulated transcription (AUX/IAA and ARF classes of genes) were also observed to change in expression, suggesting a mechanism for signal transduction at the start of ripening.
The delay in ripening observed in MADS8/9-suppressed apples may be partly due to high auxin concentrations. We propose that, to achieve low auxin associated with fruit maturation, the auxin homeostasis is controlled in a two-pronged manner: (i) by the reduction in biosynthesis and (ii) by an increase in auxin conjugation. This is associated with the change in expression of auxin-signalling genes and the up-regulation of ripening-related genes.
In russeting of apple and pear fruit, a stiff cuticle is replaced by a more plastic periderm. Furthermore, the cell layers underlying the cuticle and the periderm represent the load-bearing structure in the fruit skin in both apple and pear.
Background and aims
Russeting in apples (Malus × domestica Borkh.) and pears (Pyrus communis L.) is a disorder of the fruit skin that results from microscopic cracks in the cuticle and the subsequent formation of a periderm. To better understand russeting, rheological properties of cuticular membranes (CM) and periderm membranes (PM) were studied from the russet-sensitive apple ‘Karmijn de Sonnaville’ and from ‘Conference’ pear.
The CM and PM were isolated enzymatically, investigated by microscopy and subjected to tensile tests, creep/relaxation tests and to stepwise creep tests using a material testing machine.
The isolated CM formed a continuous polymer, whereas the PM represented a cellular structure of stacked cork cells. Tensile tests revealed higher plasticity of the hydrated PM compared with the CM, as indicated by a higher strain at the maximum force (ɛmax) and a lower modulus of elasticity (E). In apple, the maximum force (Fmax) was higher in the CM than in the PM but in pear the higher Fmax value was found for the PM. In specimens obtained from the CM : PM transition zone, the weak point in apple was found to be at the CM : PM borderline but in pear it was within the CM. In both apple and pear, creep/relaxation tests revealed elastic strain, creep strain, viscoelastic strain and viscous strain components in both the PM and CM. For any particular force, strains were always greater in the PM than in the CM and were also greater in pear than in apple. The ɛmax and Fmax values of the CM and PM were lower than those of non-russeted and russeted whole-fruit skin segments, which included adhering tissue.
In russeting, stiff CM are replaced by more plastic PM. Further, the cell layers underlying the CM and PM represent the load-bearing structure in the fruit skin in apple and pear.
cuticular membrane; fracture; fruit skin; mechanical properties; rheology; russet; strain
Nitric oxide (NO) is a plant signal contributing to plant stress responses and development. We here review some of the key advances in this field but also highlight certain key aspects of plant NO biology that require further attention.
Background and aims
After a series of seminal works during the last decade of the 20th century, nitric oxide (NO) is now firmly placed in the pantheon of plant signals. Nitric oxide acts in plant–microbe interactions, responses to abiotic stress, stomatal regulation and a range of developmental processes. By considering the recent advances in plant NO biology, this review will highlight certain key aspects that require further attention.
Scope and conclusions
The following questions will be considered. While cytosolic nitrate reductase is an important source of NO, the contributions of other mechanisms, including a poorly defined arginine oxidizing activity, need to be characterized at the molecular level. Other oxidative pathways utilizing polyamine and hydroxylamine also need further attention. Nitric oxide action is dependent on its concentration and spatial generation patterns. However, no single technology currently available is able to provide accurate in planta measurements of spatio-temporal patterns of NO production. It is also the case that pharmaceutical NO donors are used in studies, sometimes with little consideration of the kinetics of NO production. We here include in planta assessments of NO production from diethylamine nitric oxide, S-nitrosoglutathione and sodium nitroprusside following infiltration of tobacco leaves, which could aid workers in their experiments. Further, based on current data it is difficult to define a bespoke plant NO signalling pathway, but rather NO appears to act as a modifier of other signalling pathways. Thus, early reports that NO signalling involves cGMP—as in animal systems—require revisiting. Finally, as plants are exposed to NO from a number of external sources, investigations into the control of NO scavenging by such as non-symbiotic haemoglobins and other sinks for NO should feature more highly. By crystallizing these questions the authors encourage their resolution through the concerted efforts of the plant NO community.
Using sensitive and real-time detection of volatiles from plants with state-of-the-art laser based- and mass spectrometry-based methods many, hypotheses can be tested, revealing the role of the key elements in signalling and action mechanisms in plants.
Trace gas monitoring plays an important role in many areas of life sciences ranging from agrotechnology, microbiology, molecular biology, physiology, and phytopathology. In plants, many processes can be followed by their low-concentration gas emission, for compounds such as ethylene, nitric oxide, ethanol or other volatile organic compounds (VOCs). For this, numerous gas-sensing devices are currently available based on various methods. Among them are the online trace gas detection methods; these have attracted much interest in recent years. Laser-based infrared spectroscopy and proton transfer reaction mass spectrometry are the two most widely used methods, thanks to their high sensitivity at the single part per billion level and their response time of seconds. This paper starts with a short description of each method and presents performances within a wide variety of biological applications. Using these methods, the dynamics of trace gases for ethylene, nitric oxide and other VOCs released by plants under different conditions are recorded and analysed under natural conditions. In this way many hypotheses can be tested, revealing the role of the key elements in signalling and action mechanisms in plants.
Ethylene; laser-based detection; nitric oxide; plant volatiles; proton transfer reaction mass spectrometry; real-time emission; trace gas detection; volatile organic compounds
In this point of view paper, we argue that the monocot genus Hemerocallis (daylily) satisfies multiple criteria for selection as a ‘new model organism’ for intensive biological investigation. We discuss its important and interesting attributes at the biological, horticultural and medicinal levels. These include an intriguing self-incompatibility system, a sophisticated mechanism for flower bud opening and programmed floral death, and a long history of use by man as a vegetable, ornamental and medicinal plant. We examine the potential for modern technical developments to transform Hemerocallis into a valuable model plant.
Genetic model organisms have revolutionized science, and today, with the rapid advances in technology, there is significant potential to launch many more plant species towards model status. However, these new model organisms will have to be carefully selected. Here, we argue that Hemerocallis (daylily) satisfies multiple criteria for selection and deserves serious consideration as a subject of intensive biological investigation. Several attributes of the genus are of great biological interest. These include the strict control of flower opening and, within a short period, the precisely regulated floral death by a programmed cell death system. The self-incompatibility system in Hemerocallis is also noteworthy and deserves more attention. Importantly, the genus is widely cultivated for food, medicinal value and ornamental interest. Hemerocallis has considerable potential as a ‘nutraceutical’ food plant and the source of new compounds with biomedical activity. The genus has also been embraced by ornamental plant breeders and the extraordinary morphological diversity of hybrid cultivars, produced within a relatively short time by amateur enthusiasts, is an exceptional resource for botanical and genetic studies. We explore these points in detail, explaining the reasons why this genus has considerable value—both academic and socio-economic—and deserves new resources devoted to its exploration as a model. Its impact as a future model will be enhanced by its amenability to cultivation in laboratory and field conditions. In addition, established methods for various tissue and cell culture systems as well as transformation will permit maximum exploitation of this genus by science.
Asparagales; daylily; flower opening; medicinal plant; model organism; programmed cell death; self-incompatibility.
Here we discuss opportunities for system-wide analysis of plant volatiles provided by the implementation of non-supervised data processing. We illustrate the value of such approaches by presenting recent findings on wild tobacco volatile emissions using two-dimensional gas chromatography.
Plant volatile organic compound (VOC) production requires a complex network of biochemical pathways, which, although well mapped from a biochemical point of view, remains only partly understood with regard to its physiological and genetic regulation. Additionally, although analytical procedures for plant VOC measurement have become increasingly faster and more sensitive in recent years, pinpointing relevant shifts in VOC production from the thousands of molecular fragments that are generated by modern mass spectrometer instruments remains challenging. Here we discuss novel opportunities for system-wide analysis provided by the implementation of non-targeted data processing and multivariate statistics in VOC analysis. We illustrate the value of implementing non-targeted data processing with examples of recent findings from our group on the interactive control exerted by salivary components of a lepidopteran herbivore, Manduca sexta, on herbivory-induced VOC emissions in the wild tobacco Nicotiana attenuata. Finally, we briefly discuss the use of multi-platform data integration for probing the nature of metabolic and regulatory systems underlying VOC emissions.
Herbivory; metabolomics; Nicotiana attenuata; plant volatiles; two-dimensional gas chromatography; untargeted analysis.
During pollination and pollen tube growth, DNA damage is induced in the male gametes. However, detailed data on the DNA damage response is not available. Our results indicate that the generative cells of plants recognize and manage genomic lesions during pollen tube growth. In particular, we have shown that cell cycle progression of pollen mitosis II is strictly regulated by the spindle assembly checkpoint in response to genomic lesions.
Male gametophytes of plants are exposed to environmental stress and mutagenic agents during the double fertilization process and therefore need to repair the DNA damage in order to transmit the genomic information to the next generation. However, the DNA damage response in male gametes is still unclear. In the present study, we analysed the response to DNA damage in the generative cells of Cyrtanthus mackenii during pollen tube growth. A carbon ion beam, which can induce DNA double-strand breaks (DSBs), was used to irradiate the bicellular pollen, and then the irradiated pollen grains were cultured in a liquid culture medium. The male gametes were isolated from the cultured pollen tubes and used for immunofluorescence analysis. Although inhibitory effects on pollen tube growth were not observed after irradiation, sperm cell formation decreased significantly after high-dose irradiation. After high-dose irradiation, the cell cycle progression of generative cells was arrested at metaphase in pollen mitosis II, and phosphorylated H2AX (γH2AX) foci, an indicator of DSBs, were detected in the majority of the arrested cells. However, these foci were not detected in cells that were past metaphase. Cell cycle progression in irradiated generative cells is regulated by the spindle assembly checkpoint, and modification of the histones surrounding the DSBs was confirmed. These results indicate that during pollen tube growth generative cells can recognize and manage genomic lesions using DNA damage response pathways. In addition, the number of generative cells with γH2AX foci decreased with culture prolongation, suggesting that the DSBs in the generative cells are repaired.
DNA double-strand break; DNA repair; generative cell; heavy ion beam; pollen; spindle assembly checkpoint; sperm cell
Little is known about how herbivore-induced plant volatiles affect omnivorous predators. Here we show that the key predator Anthocoris nemorum is differentially attracted to three Salix clones when these are damaged by the detrimental blue willow beetle (Phratora vulgatissima). At least two volatile plant compounds were induced by the herbivore, and these were antennal active in the predator. The results elucidate how plants may recruit omnivorous predators when damaged. These findings could be utilized in crop breeding for increased resistance against herbivores.
While carnivores are known to be attracted to herbivore-induced plant volatiles, little is known about how such volatiles may affect the behaviour of omnivorous predators that may use both plants and herbivores as food. Here, we examine how systemically produced plant volatiles, in response to local herbivore damage, differentially attract a key omnivorous predator, Anthocoris nemorum (Heteroptera: Anthocoridae), to single clones of three species of Salix: S. viminalis, S. dasyclados and S. cinerea. The profiles of the plant volatiles produced were found to vary among Salix clones and between herbivore-damaged and intact plants. Anthocoris nemorum was attracted to the volatiles released from undamaged plants of all three species, but most strongly to a native S. cinerea clone. Plants damaged by the herbivorous leaf beetle Phratora vulgatissima (Coleoptera: Chrysomelidae) were generally more attractive than undamaged plants, with A. nemorum responding to systemic changes in the damaged plants where the experimental design specifically excluded volatiles released from the actual site of damage. When comparing damaged plants, the S. dasyclados clone was more attractive to A. nemorum than the S. viminalis clone—a somewhat surprising result since this Salix clone is considered relatively resistant to P. vulgatissima, and hence offers a limited amount of prey. Our experiments highlight that both constitutive and induced plant volatiles play a role in omnivore attraction, and this emphasizes the importance of considering odours of released volatiles when cropping and breeding Salix for increased resistance to herbivores.
Biocontrol; biological control; blue willow beetle; common flowerbug; E-4,8-dimethyl-1,3,7-nonatriene; GC electro-antennogram; Z-3-hexenyl acetate; short rotation coppice
We developed microsatellites for Manilkara multifida for future conservation genetics studies. M. multifida is a tropical tree that is endemic to Brazil which is currently restricted to fragmented landscapes. Our analysis indicated that all eight microsatellites are promising for assessing population genetics questions in this species.
Manilkara multifida is a tropical tree that is endemic to the Atlantic forests of southern Bahia, Brazil. Currently, populations of this species are restricted to fragmented landscapes that are susceptible to anthropogenic disturbances. Considering this issue, and that there is no genetic information available for this endangered species, we developed microsatellite markers for M. multifida to provide resources for future conservation genetics studies. Using an enriched genomic library, we isolated eight polymorphic microsatellite loci and optimized the amplification conditions for M. multifida. For each locus, we estimated the number of alleles, HE and HO, paternity exclusion Q, individual identity I and fixation index F, and examined the presence of null alleles. The mean number of alleles was 11.9, and the heterozygosity was high at all loci (average HE = 0.809 and HO = 0.777). The combined values for both paternity exclusion and individual identity were Q = 0.9959 and I = 5.45 × 10–11, respectively. No evidence of null alleles was detected. The results of our analysis indicated that all eight microsatellites are promising for assessing questions involving inbreeding, gene flow, co-ancestry and mating patterns in M. multifida.
Conservation; fragmentation; molecular ecology; molecular marker; PCR; population genetics; SSR; tropical rainforest.
Submergence inhibits photosynthesis by terrestrial wetland plants, but less so in species that possess leaf gas films when submerged. Floodwaters are often supersaturated with dissolved CO2 enabling photosynthesis by submerged terrestrial plants, although rates remain well-below those in air. This important adaptation that enhances survival in submerged conditions is reviewed.
Background and aims
Dendrobium hookerianum is a rare and threatened epiphytic orchid of northeast India. Prospects for conservation would be strengthened by developing an in vitro method for mass propagation. Seeds are minute and difficult to use directly in the field for this purpose, being non-endospermous with a low nutrient content and dependent on a specific fungus for germination and early seedling development. Although produced in large numbers (2–3 million per capsule), <5 % germinate naturally in the wild. Our objective was to develop a rapid and successful method for in vitro propagation based on an initial in vitro asymbiotic seed germination step that achieved high percentages.
Effects of four different media, i.e. (i) Murashige and Skoog (MS), (ii) Mitra et al., (iii) Knudson (KC) and (iv) Gamborg et al. (B5), were evaluated for large-scale multiplication by asymbiotic seed germination. Seedling leaf number, shoot number, shoot length, root number and root length were scored. After 7–8 months, large numbers of well-rooted plantlets were transferred to a glasshouse in thermocol pots containing compost. Six different composts based on broken brick and charcoal were compared for their ability to support further development over 90 days of hardening.
The fastest and highest percentage seed germination was achieved using MS medium. Seeds on MS medium germinated in 3–4 weeks compared with 7–8 weeks on B5 medium. Seedling development was also superior on MS medium. The inclusion of plant growth regulators was unnecessary. Compost comprising broken brick and charcoal with an upper layer of moss was found to be the most suitable for the survival of transferred plantlets. Ninety per cent survival of plantlets was achieved 90 days after transfer to a glasshouse.
The use of MS culture medium is well suited for the mass multiplication of D. hookerianum plants intended for re-introducing this threatened orchid into the wild.
Molecular genetic diversity and population structure analysis were used to clarify the controversial botanical classification of Stylosanthes guianensis. The accessions were clustered in nine groups, each of which was mainly composed of only one of the four botanical varieties.
Background and aims
The botanical classification of Stylosanthes guianensis is controversial, and few studies have used molecular markers to analyse this species. We used microsatellite markers to study the genetic diversity and population structure of S. guianensis and compare our results with the current infraspecific botanical classification.
A representative sample from the S. guianensis Brazilian germplasm collection (150 accessions) was analysed using 20 microsatellite loci. A model-based Bayesian approach implemented in the software STRUCTURE was used to assign accessions into clusters. A dendrogram was constructed based on Roger's genetic distances.
The number of alleles per locus varied from 2 to 11, with an average of 4.7. The observed (HO) and expected (HE) heterozygosity values varied from 0 to 0.58 (mean of 0.18) and from 0.04 to 0.83 (mean of 0.55), respectively. Nine groups were assembled in STRUCTURE, and these groups were consistent with clusters inferred from the genetic distances and taxonomic varieties described for S. guianensis. The GST among the nine groups was 0.46.
The low HO and the GST values observed are in agreement with the outcrossing rate (26 %) estimated for this species. The data indicate a high genetic diversity among and within the botanical varieties and suggest that microsatellite-based information can be combined with classical taxonomy to elucidate infraspecific levels.
The genome size and organization of the important medicinal plant Catharanthus roseus is shown to correspond to 1C = 0.76 pg (~738 Mbps) and 2n = 16 chromosomes. The data provide a sound basis for future studies including cytogenetic mapping, genomics and breeding.
Background and aims
Catharanthus roseus is a highly valuable medicinal plant producing several terpenoid indole alkaloids (TIAs) with pharmaceutical applications, including the anticancer agents vinblastine and vincristine. Due to the interest in its TIAs, C. roseus is one of the most extensively studied medicinal plants and has become a model species for the study of plant secondary metabolism. However, very little is known about the cytogenetics and genome size of this species, in spite of their importance for breeding programmes, TIA genetics and emerging genomic research. Therefore, the present paper provides a karyotype description and fluorescence in situ hybridization (FISH) data for C. roseus, as well as a rigorous characterization of its genome size.
The organization of C. roseus chromosomes was characterized using several DNA/chromatin staining techniques and FISH of rDNA. Genome size was investigated by flow cytometry using an optimized methodology.
The C. roseus full chromosome complement of 2n = 16 includes two metacentric, four subtelocentric and two telocentric chromosome pairs, with the presence of a single nucleolus organizer region in chromosome 6. An easy and reliable flow cytometry protocol for nuclear genome analysis of C. roseus was optimized, and the C-value of this species was estimated to be 1C = 0.76 pg, corresponding to 738 Mbp.
The organization and size of the C. roseus genome were characterized, providing an important basis for future studies of this important medicinal species, including further cytogenetic mapping, genomics, TIA genetics and breeding programmes.
The ating evidence suggests non-symbiotic hemoglobins affect hormone responses by scavenging NO. Auxin, jasmonic acid, salicylic acid, ethylene and abscisic acid have altered responses when hemoglobins are expressed. Non-symbiotic hemoglobin is a factor during plant development, biotic and abiotic stress.
Background and aims
Non-symbiotic haemoglobins have been an active research topic for over 30 years, during which time a considerable portfolio of knowledge has accumulated relative to their chemical and molecular properties, and their presence and mode of induction in plants. While progress has been made towards understanding their physiological role, there remain a number of unanswered questions with respect to their biological function. This review attempts to update recent progress in this area and to introduce a hypothesis as to how non-symbiotic haemoglobins might participate in regulating hormone signal transduction.
Advances have been made towards understanding the structural nuances that explain some of the differences in ligand association characteristics of class 1 and class 2 non-symbiotic haemoglobins. Non-symbiotic haemoglobins have been found to function in seed development and germination, flowering, root development and differentiation, abiotic stress responses, pathogen invasion and symbiotic bacterial associations. Microarray analyses under various stress conditions yield uneven results relative to non-symbiotic haemoglobin expression. Increasing evidence of the role of nitric oxide (NO) in hormone responses and the known involvement of non-symbiotic haemoglobins in scavenging NO provide opportunities for fruitful research, particularly at the cellular level.
Circumstantial evidence suggests that non-symbiotic haemoglobins may have a critical function in the signal transduction pathways of auxin, ethylene, jasmonic acid, salicylic acid, cytokinin and abscisic acid. There is a strong need for research on haemoglobin gene expression at the cellular level relative to hormone signal transduction.
Filter cubes made with machine-vision dichroic filters and illuminated with a royal blue light emitting diode can be used to produce an epifluorescent digital camera attachment that improves whole organism green fluorescent protein (GFP) photography. Mean pixel intensity responds linearly to purified GFP titration.
Background and aims
Studies have shown that levels of green fluorescent protein (GFP) leaf surface fluorescence are directly proportional to GFP soluble protein concentration in transgenic plants. However, instruments that measure GFP surface fluorescence are expensive. The goal of this investigation was to develop techniques with consumer digital cameras to analyse GFP surface fluorescence in transgenic plants.
Inexpensive filter cubes containing machine vision dichroic filters and illuminated with blue light-emitting diodes (LED) were designed to attach to digital single-lens reflex (SLR) camera macro lenses. The apparatus was tested on purified enhanced GFP, and on wild-type and GFP-expressing arabidopsis grown autotrophically and heterotrophically.
Spectrum analysis showed that the apparatus illuminates specimens with wavelengths between ∼450 and ∼500 nm, and detects fluorescence between ∼510 and ∼595 nm. Epifluorescent photographs taken with SLR digital cameras were able to detect red-shifted GFP fluorescence in Arabidopsis thaliana leaves and cotyledons of pot-grown plants, as well as roots, hypocotyls and cotyledons of etiolated and light-grown plants grown heterotrophically. Green fluorescent protein fluorescence was detected primarily in the green channel of the raw image files. Studies with purified GFP produced linear responses to both protein surface density and exposure time (H0: β (slope) = 0 mean counts per pixel (ng s mm−2)−1, r2 > 0.994, n = 31, P < 1.75 × 10−29).
Epifluorescent digital photographs taken with complementary metal-oxide-semiconductor and charge-coupled device SLR cameras can be used to analyse red-shifted GFP surface fluorescence using visible blue light. This detection device can be constructed with inexpensive commercially available materials, thus increasing the accessibility of whole-organism GFP expression analysis to research laboratories and teaching institutions with small budgets.
This paper compiles and discusses all currently available nuclear genome size data for red algae in relation to their most recent taxonomic classification.
Background and aims
The red algae are an evolutionarily ancient group of predominantly marine organisms with an estimated 6000 species. Consensus higher-level molecular phylogenies support a basal split between the unicellular Cyanidiophytina and morphologically diverse Rhodophytina, the later subphylum containing most red algal species. The Rhodophytina is divided into six classes, of which five represent early diverging lineages of generally uninucleate species, whose evolutionary relationships are poorly resolved. The remaining species compose the large (27 currently recognized orders), morphologically diverse and typically multinucleate Florideophyceae. Nuclear DNA content estimates have been published for <1 % of the described red algae. The present investigation summarizes the state of our knowledge and expands our coverage of DNA content information from 196 isolates of red algae.
The DNA-localizing fluorochrome DAPI (4′,6-diamidino-2-phenylindole) and RBC (chicken erythrocytes) standards were used to estimate 2C values with static microspectrophotometry.
Nuclear DNA contents are reported for 196 isolates of red algae, almost doubling the number of estimates available for these organisms. Present results also confirm the reported DNA content range of 0.1–2.8 pg, with species of Ceramiales, Nemaliales and Palmariales containing apparently polyploid genomes with 2C = 2.8, 2.3 and 2.8 pg, respectively.
Early diverging red algal lineages are characterized by relatively small 2C DNA contents while a wide range of 2C values is found within the derived Florideophyceae. An overall correlation between phylogenetic placement and 2C DNA content is not apparent; however, genome size data are available for only a small portion of red algae. Current data do support polyploidy and aneuploidy as pervasive features of red algal genome evolution.
Little is known about the genome of Anthurium other than chromosome observations, which frequently indicate supernumerary (“B”) chromosomes. New genome size estimates for 34 species and nine cultivars presented here provide insights into genome organization and evolution in this very large genus.
Background and aims
Anthurium is an important horticultural crop from the family Araceae, order Alismatales, a lineage considered to have diverged from other monocots prior to the cereals. Genome size and its distribution in Anthurium were investigated to gain a basic understanding of genome organization in this large genus and to forge a firm foundation for advancement of molecular approaches for the study of Anthurium. Currently, genome size estimates have been reported for only two Anthurium samples.
Bulk nuclear DNA content estimates were obtained by flow cell cytometry using leaf tissue collected from Anthurium species of different subgeneric groups and from commercial cultivars. The most current and well-supported topology of subgeneric, sectional relationships was applied to present genome size estimates in the context of reported chromosome counts, karyotypes, putative phylogenetic relationships, observed phenotypes and pedigree.
Genome size estimates based on bulk nuclear DNA content for 77 accessions representing 34 species and 9 cultivars were obtained, including initial estimates for 33 Anthurium species, and both the smallest (Anthurium obtusum; Tetraspermium) and largest (Anthurium roseospadix; Calomystrium) Anthurium genome sizes reported to date. Genome size did not distinguish any subgeneric section, but ranged 5-fold (4.42–20.83 pg/2 C) despite consistent 2N= 30 chromosome counts. Intraspecies genome size variation >20 % is reported for Anthurium ravenii, A. watermaliense and A. gracile.
Genome size estimates for Anthurium species spanning 13 recognized subgeneric sections indicate that genome size does not generally correlate with chromosome count or phylogenetic relationships. Mechanisms of genome expansion and contraction, including amplification and reduction of repetitive elements, polyploidy, chromosome reorganization/loss, may be involved in genome evolution in Anthurium as in other species. The new information on Anthurium genome sizes provides a platform for molecular studies supporting further research on genome evolution as well as cultivar development.
Ecological traits of the circumboreal plant Viburnum opulus were examined to improve understanding of the variation of populations occurring in the same biome but on different continents. Seedling development/emergence is shown to be highly similar but some degree of variation was present in other traits, among populations.
Background and aims
Temperate forests are disjunct in the Northern Hemisphere, having become fragmented from the earlier widespread (Tertiary) boreotropical forest. We asked ‘What are the contemporary patterns of population variation in ecological traits of a Tertiary relict in a macroecological context?’. This issue underpins our understanding of variation in populations occurring in the same biome but on different continents.
We examined characters associated with root and shoot emergences among populations of Viburnum opulus in temperate forests of Asia, North America and Europe. This species has complex seedling emergence extending over several years and requiring various temperature cues.
Populations varied in germination responses and clustered into groups that were only partly related to varietal status. Whereas roots (at warm temperatures) and shoots (following a cold period) simultaneously emerged from seeds of all populations when simulated dispersal occurred in winter, they were delayed in some populations when dispersal occurred in summer.
Viburnum opulus populations, some separated by 10 300 km, showed high similarity in seedling development and in germination phenology, and we suggest that stabilizing selection has played a key role in maintaining similar dormancy mechanisms. Nevertheless, there was some degree of variation in other germination characters, suggesting local adaptation.
Grapevines growing in Australia suffer from high temperatures which have major effects on photosynthesis and transpiration. To learn more, gas exchange was measured over several seasons and then modelled across temperatures from 20 to 45°C and validated with independent data.
Background and aims
Grapevines growing in Australia are often exposed to very high temperatures and the question of how the gas exchange processes adjust to these conditions is not well understood. The aim was to develop a model of photosynthesis and transpiration in relation to temperature to quantify the impact of the growing conditions on vine performance.
Leaf gas exchange was measured along the grapevine shoots in accordance with their growth and development over several growing seasons. Using a general linear statistical modelling approach, photosynthesis and transpiration were modelled against leaf temperature separated into bands and the model parameters and coefficients applied to independent datasets to validate the model.
Photosynthesis, transpiration and stomatal conductance varied along the shoot, with early emerging leaves having the highest rates, but these declined as later emerging leaves increased their gas exchange capacities in accordance with development. The general linear modelling approach applied to these data revealed that photosynthesis at each temperature was additively dependent on stomatal conductance, internal CO2 concentration and photon flux density. The temperature-dependent coefficients for these parameters applied to other datasets gave a predicted rate of photosynthesis that was linearly related to the measured rates, with a 1 : 1 slope. Temperature-dependent transpiration was multiplicatively related to stomatal conductance and the leaf to air vapour pressure deficit and applying the coefficients also showed a highly linear relationship, with a 1 : 1 slope between measured and modelled rates, when applied to independent datasets.
The models developed for the grapevines were relatively simple but accounted for much of the seasonal variation in photosynthesis and transpiration. The goodness of fit in each case demonstrated that explicitly selecting leaf temperature as a model parameter, rather than including temperature intrinsically as is usually done in more complex models, was warranted.
Austrobaileya has long served as a model for ancient angiosperm pollen structure. Its pollen germination is relatively rapid and requires < 10 % of the progamic phase. Extensive evidence suggests pollen germination underwent acceleration early in angiosperm history.
Background and aims
The pollination to fertilization process (progamic phase) is thought to have become greatly abbreviated with the origin of flowering plants. In order to understand what developmental mechanisms enabled the speeding of fertilization, comparative data are needed from across the group, especially from early-divergent lineages. I studied the pollen germination process of Austrobaileya scandens, a perennial vine endemic to the Wet Tropics area of northeastern Queensland, Australia, and a member of the ancient angiosperm lineage, Austrobaileyales.
I used in vivo and in vitro hand pollinations and timed collections to study development from late pollen maturation to just after germination. Then I compared the contribution of pollen germination timing to progamic phase duration in 131 angiosperm species (65 families).
Mature pollen of Austrobaileya was bicellular, starchless and moderately dehydrated—water content was 31.5 % by weight and volume increased by 57.9 % upon hydration. A callose layer in the inner intine appeared only after pollination. In vivo pollen germination followed a logarithmic curve, rising from 28 % at 1 hour after pollination (hap) to 97 % at 12 hap (R2 = 0.98). Sufficient pollen germination to fertilize all ovules was predicted to have occurred within 62 min. Across angiosperms, pollen germination ranged from 1 min to >60 h long and required 8.3 ± 9.8 % of the total duration of the progamic phase.
Pollen of Austrobaileya has many plesiomorphic features that are thought to prolong germination. Yet its germination is quite fast for species with desiccation-tolerant pollen (range: <1 to 60 h). Austrobaileya and other early-divergent angiosperms have relatively rapid pollen germination and short progamic phases, comparable to those of many insect-pollinated monocots and eudicots. These results suggest that both the pollen germination and pollen tube growth periods were marked by acceleration of developmental processes early in angiosperm history.
Cells of embryonic axes in recalcitrant horse chestnut seeds are associated with a high water content and functionally preserved vacuoles with active aquaporins and invertase. These vacuoles function to the support rapid cell elongation of the hypocotyl that results in radicle emergence and thus the start of visible germination.
Backgrounds and aims
In tropical recalcitrant seeds, their rapid transition from shedding to germination at high hydration level is of physiological interest but difficult to study because of the time constraint. In recalcitrant horse chestnut seeds produced in central Russia, this transition is much longer and extends through dormancy and dormancy release. This extended time period permits studies of the water relations in embryonic axes during the long recalcitrant period in terms of vacuolar status and water transport.
Horse chestnut (Aesculus hippocastanum) seeds sampled in Moscow were stratified in cold wet sand for 4 months. Vacuole presence and development in embryonic axes were examined by vital staining, light and electron microscopy. Aquaporins and vacuolar H+-ATPase were identified immunochemically. Water channel operation was tested by water inflow rate. Vacuolar acid invertase was estimated in terms of activity and electrophoretic properties.
Throughout the long recalcitrant period after seed shedding, cells of embryonic axes maintained active vacuoles and a high water content. Preservation of enzyme machinery in vacuoles was evident from retention of invertase activity, substrate specificity, molecular mass and subunit composition. Plasmalemma and tonoplast aquaporins and the E subunit of vacuolar H+-ATPase were also present. In non-dormant seeds prior to growth initiation, vacuoles enlarged at first in hypocotyls, and then in radicles, with their biogenesis being similar. Vacuolation was accompanied by increasing invertase activity, leading to sugar accumulation and active osmotic functioning. After growth initiation, vacuole enlargement was favoured by enhanced water inflow through water channels formed by aquaporins.
Maintenance of high water content and desiccation sensitivity, as well as preservation of active vacuoles in embryonic axes after shedding, can be considered a specific feature of recalcitrant seeds, overlooked when studying tropical recalcitrants due to the short duration. The retained physiological activity of vacuoles allows them to function rapidly as dormancy is lost and when external conditions permit. Cell vacuolation precedes cell elongation in both hypocotyl and radicle, and provides impetus for rapid germination.
The paper describes the functional analysis of a class C heat shock transcription factor from rice (Oryza sativa). OsHsfC1b is shown to play a role in ABA-mediated salt stress tolerance and is required for plant growth under non-stress conditions.
Background and aims
Salt stress leads to attenuated growth and productivity in rice. Transcription factors like heat shock factors (HSFs) represent central regulators of stress adaptation. Heat shock factors of the classes A and B are well established as regulators of thermal and non-thermal stress responses in plants; however, the role of class C HSFs is unknown. Here we characterized the function of the OsHsfC1b (Os01g53220) transcription factor from rice.
We analysed the expression of OsHsfC1b in the rice japonica cultivars Dongjin and Nipponbare exposed to salt stress as well as after mannitol, abscisic acid (ABA) and H2O2 treatment. For functional characterization of OsHsfC1b, we analysed the physiological response of a T-DNA insertion line (hsfc1b) and two artificial micro-RNA (amiRNA) knock-down lines to salt, mannitol and ABA treatment. In addition, we quantified the expression of small Heat Shock Protein (sHSP) genes and those related to signalling and ion homeostasis by quantitative real-time polymerase chain reaction in roots exposed to salt. The subcellular localization of OsHsfC1b protein fused to green fluorescent protein (GFP) was determined in Arabidopsis mesophyll cell protoplasts.
Expression of OsHsfC1b was induced by salt, mannitol and ABA, but not by H2O2. Impaired function of OsHsfC1b in the hsfc1b mutant and the amiRNA lines led to decreased salt and osmotic stress tolerance, increased sensitivity to ABA, and temporal misregulation of salt-responsive genes involved in signalling and ion homeostasis. Furthermore, sHSP genes showed enhanced expression in knock-down plants under salt stress. We observed retarded growth of hsfc1b and knock-down lines in comparison with control plants under non-stress conditions. Transient expression of OsHsfC1b fused to GFP in protoplasts revealed nuclear localization of the transcription factor.
OsHsfC1b plays a role in ABA-mediated salt stress tolerance in rice. Furthermore, OsHsfC1b is involved in the response to osmotic stress and is required for plant growth under non-stress conditions.
The paper uses modified population viability models and spatial structure via block analysis to assess population demography of Trillium recurvatum a clonal understory plant. The population is expanding, a likely outcome of the relatively high proportion of juvenile and non-flowering adult ramets and fast-replicating non-flowering adults. Further work is needed to elucidate the relative contributions of clonal vs seed recruitment to genetic structure and demography.
Background and aims
Understanding the demography of long-lived clonal herbs, with their extreme modularity, requires knowledge of both their short- and long-term survival and ramet growth patterns. The primary objective of this study was to understand the dynamics of a clonal forest herb, Trillium recurvatum, by examining temporal and small-scale demographic patterns. We hypothesized: (i) there would be more variability in the juvenile age class compared with non-flowering adult and flowering adult classes due to year-to-year fluctuations in recruitment; (ii) rates of population growth (λ) and increase (r) would be highest in non-flowering ramets due to a combination of transitions from the juvenile stage and reversions from flowering adults; and (iii) inter-ramet distances would be most variable between flowering and juvenile ramets due to a combination of clonal growth, seed dispersal by ants and ramet death over time.
Census data were collected on the total number of stems in the population from 1990 to 2007, and placed within one of three life stages (juvenile, three-leaf non-flowering and three-leaf flowering). Modified population viability equations were used to assess temporal population viability, and spatial structure was assessed using block krigging. Correlations were performed using current and prior season weather to current population demography.
The first hypothesis was rejected. The second hypothesis was supported: population increase (r) and growth rate (λ) were highest in non-flowering ramets. Finally, the third hypothesis was rejected: there was no apparent density dependence within this population of Trillium and no apparent spatial structure among life stages.
Overall population density fluctuated over time, possibly due to storms that move soil, and prior year's temperature and precipitation. However, density remained at some dynamic stable level. The juvenile age class had greater variability for the duration of this study and population growth rate was greatest for non-flowering ramets.
This paper show that inter-subspecies hybridization among certain Vigna unguiculata subspecies, occurred during the course of evolution. This has affected several regions of the genome and is interfering with the dependable assessment of sub-species relationships using single (rRNA regions) or multilocus markers.
Background and aims
Intra-species hybridization and incompletely homogenized ribosomal RNA repeat units have earlier been reported in 21 accessions of Vigna unguiculata from six subspecies using internal transcribed spacer (ITS) and 5S intergenic spacer (IGS) analyses. However, the relationships among these accessions were not clear from these analyses. We therefore assessed intra-species hybridization in the same set of accessions.
Arbitrarily primed polymerase chain reaction (AP-PCR) analysis was carried out using 12 primers. The PCR products were resolved on agarose gels and the DNA fragments were scored manually. Genetic relationships were inferred by TREECON software using unweighted paired group method with arithmetic averages (UPGMA) cluster analysis evaluated by bootstrapping and compared with previous analyses based on ITS and 5S IGS.
A total of 202 (86 %) fragments were found to be polymorphic and used for generating a genetic distance matrix. Twenty-one V. unguiculata accessions were grouped into three main clusters. The cultivated subspecies (var. unguiculata) and most of its wild progenitors (var. spontanea) were placed in cluster I along with ssp. pubescens and ssp. stenophylla. Whereas var. spontanea were grouped with ssp. alba and ssp. tenuis accessions in cluster II, ssp. alba and ssp. baoulensis were included in cluster III. Close affinities of ssp. unguiculata, ssp. alba and ssp. tenuis suggested inter-subspecies hybridization.
Multi-locus AP-PCR analysis reveals that intra-species hybridization is prevalent among V. unguiculata subspecies and suggests that grouping of accessions from two different subspecies is not solely due to the similarity in the ITS and 5S IGS regions but also due to other regions of the genome.