Differences between Fragaria clones by fourth derivative room temperature spectroscopy of intact leaves

Clones of Fragaria were characterized by the attenuance (absorption) spectra of their photosynthetic apparatus. This method uses fourth derivative analysis of spectra of intact lamina of detached leaves at room temperature (20–25 C). Twenty-one different species and hybrids of Fragaria were examined. F. chiloensis accessions frequently have characteristic fourth derivative spectra distinct from that of other Fragaria species. These spectra are characterized by a strong band that appears to be photosystem I component Ca 693; and is correlated with a higher chlorophyll b content per unit area of leaf, and a lower chlorophyll a/b ratio.Abbreviations Chl chlorophyll - chiloensis Fragaria chiloensis - 4th deriv fourth derivative - PSI photosystem I - PSH photosystem II     

ECOLOGICAL DIFFERENTIATION IN PERENNIAL,OCTOPLOID SPECIES OF FRAGARIA

Plants of octoploid Fragaria (2n = 56) were collected from 19 natural sites supporting strawberry colonies in California, grown in a common greenhouse and analyzed for 25 quantitative traits and 2 enzyme systems. Inter-populational variation was measured by using analysis of variance and genetic identity techniques. Micro- and macro-environmental measurements were made at each site. Stepwise multiple regressions were completed on the morphological-environmental variations and the allozyme frequency-environmental variations. Clonal propagules were compared under three controlled environmental conditions. Octoploid Fragaria appears to have undergone considerable ecological differentiation. Significant amounts of inter-populational variation were observed in a number of polygenic and monogenic traits. Numerous significant correlations were found between environmental and character variations, and in many cases, only a few environmental variables were necessary to “explain” significant amounts of that variation. Inter-populational differences were also found in the abilities of plants to accumulate biomass and survive under salt, nutrient and shade stress. It has been suggested that ecological differentiation plays a minor role in determining the eco-geographical range of allopolyploids due to the effects of polyploidy on the generation of mutational and recombinational variability. Clearly, this has not been the case in Fragaria.     

Bioclimatic evaluation of geographical range in Fragaria (Rosaceae): consequences of variation in breeding system,ploidy and species age

The evaluation of the intrinsic and extrinsic forces that determine geographical range sizes and niche breadth is key to the understanding of species distributions and for informing the conservation of biodiversity. Fragaria (Rosaceae) contains the economically important cultivated strawberry (Fragaria × ananassa subsp. ananassa) and numerous wild species. Using georeferenced species records and global bioclimatic data, we describe the bioclimatic niches for 21 Fragaria spp. and investigate the relationship between their niches and geographical range size, breeding system, ploidy and time since divergence. We found no evidence of phylogenetic signal for bioclimatic niches. There was also no relationship between ploidy and geographical or bioclimatic range area, but geographical range area was significantly greater for species that were capable of self‐fertilization. In addition, we found a significant decelerating relationship between species age and geographical range area. Overall, our results suggest that Fragaria spp., although similar in morphology and life history, show high levels of divergence in bioclimatic niches and significant over‐dispersion along some bioclimatic gradients, suggesting evolutionary lability in physiology and climate tolerance. As a consequence, wild species will remain a valuable resource for cultivated strawberry sustainability, especially under changing future climate. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 176 , 99–114.     

Development and bin mapping of strawberry genic-SSRs in diploid <Emphasis Type="Italic">Fragaria</Emphasis> and their transferability across the Rosoideae subfamily

Cultivated strawberry (Fragaria × ananassa) together with other economically important genera such as Rosa (roses) and Rubus (raspberry and blackberry) belongs to the subfamily Rosoideae. There is increasing interest in the development of transferable markers to allow genome comparisons within the Rosaceae family. In this report, 122 new genic microsatellite (SSR) markers have been developed from cultivated strawberry and its diploid ancestor Fragaria vesca. More than 77% of the sequences from which the markers were developed show significant homology to known or predicted proteins and more than 92% were polymorphic among strawberry cultivars, representing valuable markers in transcribed regions of the genome. Sixty-three SSRs were polymorphic in the diploid Fragaria reference population and were bin-mapped together with another five previously reported but unmapped markers. In total, 72 loci were distributed across the seven linkage groups. In addition, the transferability of 174 Fragaria SSRs to the related Rosa and Rubus genera was investigated, ranging from 28.7% for genic-SSRs in rose to 16.1% for genomic-SSRs in raspberry. Among these markers, 33 and 16 were both localized in the diploid Fragaria reference map and cross-amplified in rose and raspberry, respectively. These results indicate that transferability of SSRs across the Rosoideae subfamily is limited. However, we have identified a set of Fragaria markers, polymorphic in the diploid reference population, which cross-amplified in both Rosa and Rubus, which represents a valuable tool for comparative mapping and genetic diversity analyses within the Rosoideae subfamily.     

Melting, glass transition, and apparent heat capacity of α-d-glucose by thermal analysis

The thermal behaviors of α-d-glucose in the melting and glass transition regions were examined utilizing the calorimetric methods of standard differential scanning calorimetry (DSC), standard temperature-modulated differential scanning calorimetry (TMDSC), quasi-isothermal temperature-modulated differential scanning calorimetry (quasi-TMDSC), and thermogravimetric analysis (TGA). The quantitative thermal analyses of experimental data of crystalline and amorphous α-d-glucose were performed based on heat capacities. The total, apparent and reversing heat capacities, and phase transitions were evaluated on heating and cooling. The melting temperature (T_m) of a crystalline carbohydrate such as α-d-glucose, shows a heating rate dependence, with the melting peak shifted to lower temperature for a lower heating rate, and with superheating of around 25 K. The superheating of crystalline α-d-glucose is observed as shifting the melting peak for higher heating rates, above the equilibrium melting temperature due to of the slow melting process. The equilibrium melting temperature and heat of fusion of crystalline α-d-glucose were estimated. Changes of reversing heat capacity evaluated by TMDSC at glass transition (T_g) of amorphous and melting process at T_m of fully crystalline α-d-glucose are similar. In both, the amorphous and crystalline phases, the same origin of heat capacity changes, in the T_g and T_m area, are attributable to molecular rotational motion. Degradation occurs simultaneously with the melting process of the crystalline phase. The stability of crystalline α-d-glucose was examined by TGA and TMDSC in the melting region, with the degradation shown to be resulting from changes of mass with temperature and time. The experimental heat capacities of fully crystalline and amorphous α-d-glucose were analyzed in reference to the solid, vibrational, and liquid heat capacities, which were approximated based on the ATHAS scheme and Data Bank.     

Conformational studies of peptides: Crystal and molecular structures of L-3,4-dehydroproline and its t-butoxycarbonyl and acetyl amide derivatives

The crystal structures of L -3,4-dehydroproline, t-butoxycarbonyl-L -3,4-dehydroproline amide, and acetyl-L -3,4-dehydroproline amide have been determined. L -3,4-Dehydroproline is orthorhombic with a = 16.756, b = 5.870, c = 5.275 Å, and Z = 4; t-butoxycarbonyl-L -3,4-dehydroproline amide is orthorhombic with a = 6.448, b = 8.602, c = 21.710 Å, and Z = 4; acetyl-L -3,4-dehydroproline amide is monoclinic with a = 4.788, b = 10.880, c = 7.785 Å, β = 105.25°, and Z = 2. The final R value for the L -3,4-dehydroproline is 0.046 based on 529 reflections; for t-butoxycarbonyl-L -3,4-dehydroproline amide, 0.050 based on 792 reflections; and for acetyl-L -3,4-dehydroproline amide, 0.058 based on 632 reflections. The structures clearly establish that the free amino acid exists in the zwitterionic form in the crystalline state. The molecular conformations of the t-Boc and acetyl derivatives consist of two planes: one involving the primary amide and the other the remaining atoms of the molecule. The acetyl-L -3,4-dehydroproline amide contains a tertiary amide bond in the cis conformation. To the best of our knowledge, this is the first example of a cis bond in an acetyl derivative of an amino acid or peptide. At variance with the previously reported proline amides, which present ? and ψ values corresponding to those of a right-handed α-helical conformation (conformation A), the t-Boc and acetyl derivatives both have ? and ψ values corresponding to a collagenlike conformation (conformation F).     

Bio-degradation and utilization of silica and quartz

Summary A mesophilic bacterium,Proteus mirabilis, which was known to be able to accumulate monomer silicate ions, a thermophilic bacterium,Bacillus caldolyticus, originating from a habitat with high silica concentrations, and a silicautilizing plant,Equisetum arvense, were all found to produce monomer silica from its polymer. The monomer silica, resulting from the mineralysis of either experimentally polymerized silica, or from quartz, is taken up byP. mirabilis cells, and also byEquisetum, which then deposits the silica again as a polymer in its stem and leaves. WithB. caldolyticus, which does not utilize the depolymerized product under the given conditions, we found that the intensity of the mineralysis depends on the growth rate of the organism.     

(Z)-3-Hexenyl-β-d-glucopyranoside in Fresh Tea Leaves as a Precursor of Green Odor

The glycoside fraction from fresh tea leaves was acetylated and separated by silica gel column chromatography.

A crystalline product was identified as (Z)-3-hexenyl-(tetra-O-acetyl)-β-d-glucopyranoside from spectrometric data which were identical with those of an authentic synthesized sample in all respects.

There are two possible processes for the formation of the greenish odor of plant materials, these being a biosynthetic process from the lipid and enzymatic hydrolysis of (Z)-3-hexenyl-β-d-glucoside.     

The development and mapping of functional markers in Fragaria and their transferability and potential for mapping in other genera

We have developed 46 primer pairs from exon sequences flanking polymorphic introns of 23 Fragaria gene sequences and one Malus sequence deposited in the EMBL database. Sequencing of a set of the PCR products amplified with the novel primer pairs in diploid Fragaria showed the products to be homologous to the sequences from which the primers were originally designed. By scoring the segregation of the 24 genes in two diploid Fragaria progenies FV × FN (F. vesca × F. nubicola F₂) and 815 × 903BC (F. vesca × F. viridis BC₁) 29 genetic loci at discrete positions on the seven linkage groups previously characterised could be mapped, bringing to 35 the total number of known function genes mapped in Fragaria. Twenty primer pairs, representing 14 genes, amplified a product of the expected size in both Malus and Prunus. To demonstrate the applicability of these gene-specific loci to comparative mapping in Rosaceae, five markers that displayed clear polymorphism between the parents of a Malus and a Prunus mapping population were selected. The markers were then scored and mapped in at least one of the two additional progenies.     

GENETICS OF SEX EXPRESSION IN FRAG ARIA SPECIES

Hermaphroditism is the normal mode of sex expression in diploid species of Fragaria (Rosaceae, 2n = 14, x = 7) with one known exception, gynodioecious F. vesca L. ssp. bracteata. The polyploid species of Fragaria are all trioecious. An extensive study involving appropriate hybridization, testcrossing, selling, and backcrossing revealed that the repression of sporangia and the inhibition of sporogenesis are controlled by a single gene (or a gene complex) with at least three alleles in the sporophytes of trioecious octoploid species (2n = 56, x = 7). A male suppressor (allele F) reduces microsporangia and represses microsporogenesis completely, but it allows normal development of the stigma, style, and ovary. A female suppressor (allele M), in the absence of modifier genes, inhibits megasporogenesis and drastically reduces the number of carpels and size of receptacles. The allele H, conferring perfect flowers at an early stage of flower organogenesis, acts as an inducer of microsporogenesis in females, but leaves both microand megasporangial development intact. At the sex locus, the F allele (femaleness) is dominant to H and M and the H allele (hermaphroditism) is dominant to M (maleness). Females are exclusively heterogametic (F/H or F/M), hermaphrodites may be homo- or heterogametic (H/H or H/M), and males are homogametic (M/M). The sex gene is expressed precisely in the genetic background of octoploid × diploid hybrids of Fragaria and their derivatives and in crosses with closely related hermaphroditic diploid Pontentilla glandulosa L. Gene dosage phenomena are absent. First generation progeny of colchi-decaploids (F/F, –/–) are exclusively female, but all generations thereafter segregate in a normal diploidized manner (1:1). Application of phytohormones alters sex expression to a limited extent.     

A UDP‐glucosyltransferase functions in both acylphloroglucinol glucoside and anthocyanin biosynthesis in strawberry (Fragaria × ananassa)

Physiologically active acylphloroglucinol (APG) glucosides were recently found in strawberry (Fragaria sp.) fruit. Although the formation of the APG aglycones has been clarified, little is known about APG glycosylation in plants. In this study we functionally characterized ripening‐related glucosyltransferase genes in Fragaria by comprehensive biochemical analyses of the encoded proteins and by a RNA interference (RNAi) approach in vivo. The allelic proteins UGT71K3a/b catalyzed the glucosylation of diverse hydroxycoumarins, naphthols and flavonoids as well as phloroglucinols, enzymatically synthesized APG aglycones and pelargonidin. Total enzymatic synthesis of APG glucosides was achieved by co‐incubation of recombinant dual functional chalcone/valerophenone synthase and UGT71K3 proteins with essential coenzyme A esters and UDP‐glucose. An APG glucoside was identified in strawberry fruit which has not yet been reported in other plants. Suppression of UGT71K3 activity in transient RNAi‐silenced fruits led to a loss of pigmentation and a substantial decrease of the levels of various APG glucosides and an anthocyanin. Metabolite analyses of transgenic fruits confirmed UGT71K3 as a UDP‐glucose:APG glucosyltransferase in planta. These results provide the foundation for the breeding of fruits with improved health benefits and for the biotechnological production of bioactive natural products.     

Stokes–Einstein violation and fragility in calcium aluminosilicate glass formers: a molecular dynamics study

A series of classical molecular dynamics simulations of calcium aluminosilicate (CAS) (CaO–Al₂O₃)_1–x(SiO₂)_x melts with varying silica content x, along the CaO/Al₂O₃ concentration ratio R = 1, have been performed for the purpose of studying (i) the evolution of fragility with silica content and (ii) the temperature and composition evolution of self-diffusion coefficients, viscosity and α-relaxation times. Our results indicate a decrease in the fragility of the CAS systems along R = 1 from calcium aluminate to pure silica. It is found that upon cooling, the Stokes–Einstein relation breaks down at a temperature significantly higher than the critical temperature of the mode-coupling theory T_C for all compositions and also above the melting point.     

Regulated Changes in the Acetylation of α-Tubulin on Lys40 during Growth and Organ Development in Fast Plants,Brassica rapa L.

Recently, we confirmed the widespread occurrence of α-tubulin acetylation on Lys⁴⁰ in angiosperms. In the present study, we found that α-tubulin acetylation is regulated in a growth stage- and organ development-dependent manner in the rapid cycling Brassica rapa, also known as Fast Plants. Organ distribution analysis showed that the proportion of acetylated α-tubulin is high in the cotyledons of young plants and in the true leaves and flowers of mature plants. A correlation between the increase in the levels of α-tubulin acetylation and the maturation of true leaves was observed. In the mature leaves, the acetylated α-tubulin showed an uneven distribution pattern, and the cells in the region of the leaf margins contained a high proportion of acetylated α-tubulin. These results indicate that α-tubulin acetylation is dynamically regulated in plant organs during development, and that it might play an important role in microtubule functioning throughout the angiosperm's life cycle.     

Assessment of the influence of different sample processing and cold storage duration on plant free proline content analyses

Introduction – A method which is widely accepted for the analysis of free proline content in plant tissues is based on the use of 3% sulfosalicylic acid as an extractant, followed by spectrophotometric quantification of a proline–ninhydrin complex in toluene. However, sample preparation and storage may influence the proline actually measured. This may give misleading or difficult to compare data. Objective and Methodology – To evaluate free proline levels fresh and frozen strawberry (Fragaria × ananassa Duch.) leaves and soybean [Glycine max (L.) Merr.] hypocotyl tissues were used. These were ground with or without liquid nitrogen and proline extracted with sulfosalicylic acid. A particular focus was the influence of plant sample cold storage duration (1, 4 and 12 weeks at ?20°C) on tissue proline levels measured. Results – The free proline content analyses, carried out in leaves of Fragaria × ananassa Duch. as well as in hypocotyls of Glycine max (L.) Merr., showed a significant influence of the sample preparation method and cold storage period. Long‐term storage of up to 12 weeks at ?20°C led to a significant increase in the measured proline in all samples analysed. Conclusion – The observed changes in proline content in plant tissue samples stored at ?20°C indicate the likelihood of the over‐estimation of the proline content if the proline analyses are delayed. Plant sample processing and cold storage duration seem to have an important influence on results of proline analyses. Therefore it is recommended that samples should be ground fresh and analysed immediately. Copyright © 2010 John Wiley & Sons, Ltd.     

Effect of High Temperature on Protein Expression in Strawberry Plants

Strawberry plants (Fragaria×ananassa Duch.) cvs. Nyoho and Toyonoka were exposed to temperatures of 20, 33, and 42 °C for 4 h, and protein patterns in leaves and flowers was analyzed by 2-dimensional polyacrylamide gel electrophoresis and immunoblotting. In leaves and flowers of both cultivars, the content of most proteins decreased, but a few new proteins appeared in response to heat stress. These heat shock proteins (Hsps) were detected in the range of 19 – 29 kDa in leaves, and 16 – 26 kDa in flowers. The intensity of a 43 kDa protein spot increased in response to heat stress in Nyoho flowers, but not in Toyonoka flowers. The peaHsp17.7 antibody recognized one band at approximately 26 kDa in leaves, and two bands at approximately 16 and 17 kDa in flowers of both cultivars. These results show that the effects of heat stress on Hsp synthesis in strawberry plants differ between plant organs and between cultivars.     

Isolation and phytotoxicity of a metabolite from Curvularia eragrostidis and characterisation of its modes of action

Digitaria sanguinalis is a widespread troublesome weed distributed all over the world. Curvularia eragrostidis QZ‐2000 is a potential candidate for biocontrol of D. sanguinalis. A phytotoxic metabolite from the culture filtrate of this fungus was extracted by ethyl acetate, isolated by bioassay‐guided column chromatography and thin layer chromatography on silica gel, characterised by ultra violet, infrared ray (IR), mass spectrometry (MS), ¹H‐nuclear magnetic resonance (NMR) and ¹³C‐NMR spectral data analyses and identified as α,β‐dehydrocurvularin. The phytotoxin significantly inhibited seed germination of D. sanguinalis from 43 to 688 μM. The EC₅₀ value of seed germination was 152 μM. The EC₅₀ values of elongation of radicle and coleoptile were 102 and 172 μM, respectively. α,β‐dehydrocurvularin caused extensive necrosis on leaves of many notorious weeds at 688 μM, while maize and soybean were insensitive to it. Therefore, α,β‐dehydrocurvularin was regarded as a non‐host‐selective phytotoxin. At concentrations of 172–688 μM, α,β‐dehydrocurvularin caused a decrease in chlorophyll content. α,β‐dehydrocurvularin had stronger impacts on chlorophyll A fluorescence, photophosphorylation and Mg²⁺‐ATPase activity at higher concentrations. These results suggest that α,β‐dehydrocurvularin affected the photosynthetic capacity. In the present study, α,β‐dehydrocurvularin significantly inhibited mitosis of root tip cells. It supports that the α,β‐dehydrocurvularin has potential for development as a natural bioherbicide.     

EST‐SSR markers from Fragaria vesca L. cv. Yellow Wonder

Fourteen microsatellite primer pairs were developed from a cDNA library of heat‐treated seedlings of Fragaria vesca cv. yellow wonder. Transferability to 13 species of Fragaria ranged from 71% in diploid species F. gracilis Losinsk., F. iinumae Makino, F. nilgerrensis Schltdl. ex J. Gay and F. nipponica Makino, to 100% in octoploid domestic strawberry and its progenitors. Polymorphism was high in polyploid Fragaria species. However, polymorphism and heterozygosity of eight EST‐SSRs (expressed sequence tag–simple sequence repeats) was low in 14 F. vesca genotypes.     

Study of Zn,Cu and Pb content in plants and contaminated soils in Sardinia

Trace elements in soils exist as components of several different fractions. We have analyzed the correlation between total and extractable (EDTA, calcium chloride and deionized water) Zn, Pb and Cu concentrations in soils and the concentration of these elements in plant leaves. Soil and plant samples have been taken from Sulcis-Iglesiente (Sardinia), an area rich in mining tailings. This has made that the concentrations of the trace element under study in soils were varied. Three plants have been studied: Dittrichia viscosa, Cistus salviifolius, and Euphorbia pithyusa subsp. cupanii. Soil samples beneath each of them at depths of 0–30 and 30–60 cm have been considered. The highest concentration of trace elements in the leaves of the studied species has been found for Zn. The calcium carbonate content and the crystalline and amorphous forms of iron in the soil have determined the concentration of metal in plant leaves. The soil concentrations that have been found with the extraction methods are uncorrelated with Pb and Cu concentrations in plants, but Zn is correlated with the fraction extracted with EDTA and calcium chloride. The concentrations of trace metals in plants are most closely related to the soil contents of CaCO₃, electrical conductivity, Fe_ox, and Fe_dc.     

Tracking the evolutionary history of polyploidy in Fragaria L. (strawberry): New insights from phylogenetic analyses of low-copy nuclear genes

Phylogenetic utility of two nuclear genes (GBSSI-2 and DHAR) was explored in genus Fragaria in order to clarify phylogenetic relationships among taxa and to elucidate the origin of the polyploid species. Orthology of the amplified products was assessed by several methods. Our results strongly suggest the loss of one GBSSI duplicated copy (GBSSI-1) in the Fragariinae subtribe. Phylogenetic analyses provided new insights into the evolutionary history of Fragaria, such as evidence supporting the presence of three main diploid genomic pools in the genus and demonstrating the occurrence of independent events of polyploidisation. In addition, the data provide evidence supporting an allopolyploid origin of the hexaploid F. moschata, and the octoploids F. chiloensis, F. iturupensis and F. virginiana. Accordingly, a new pattern summarizing our present knowledge on the Fragaria evolutionary history is proposed. Additionally, sequence analyses also revealed relaxed constraints on homoeologous copies at high ploidy level, as demonstrated by deletion events within DHAR coding sequences of some allo-octoploid haplotypes.