Divergence of chloroplast gene organization in three legumes: Pisum sativum,Vicia faba and Phaseolus vulgaris

Summary Isolated chloroplasts from Pisum sativum were found to contain at least 32 tRNA species. Hybridization of in vitro labeled, identified, chloroplast tRNAs to Pisum chloroplast DNA fragments revealed the locations of the tRNA genes on the circular chloroplast genome. Comparison of this gene map to the maps of Vicia faba and Phaseolus vulgaris showed that the chloroplast genomes of Pisum and Phaseolus are otherwise more closely related than either genome is to the chloroplast genome of Vicia. Furthermore, the results suggest how possible recombination events could be involved in the evolution of these three closely related, but divergent, chloroplast genomes.     

Bile-pigment formation from different leghaemoglobins. Methine-bridge specificity of coupled oxidation

The coupled oxidation of leghaemoglobins with O₂ and ascorbate yielded oxyleghaemoglobin in the first reaction step, and the second step was the degradation of haem characterized by an A₆₇₅ increase. Leghaemoglobins were degraded to biliverdin isomers specifically, depending on the structure of the protein. The main leghaemoglobin components of Glycine (soya bean) and Phaseolus (kidney bean) were degraded to biliverdin mixtures containing about 50% of the β-form, about 30% of the α-form and about 20% of the δ-isomer, whereas the leghaemoglobin I components of Vicia (broad bean) and Pisum (pea) were degraded almost exclusively to the β-isomer, with traces of the α-isomer. The amino acid sequences of Glycine and Phaseolus leghaemoglobins resemble each other, as do those of Vicia and Pisum. The site specificity of bile-pigment formation from leghaemoglobins can be tentatively explained by specific differences in the amino acid sequences at those regions of the polypeptide chain that are in the vicinity of the appropriate methine bridges. The ligand-binding site in different leghaemoglobins may be outlined on the basis of the present results, supposing that the haem is degraded when a reduction product of haem-bound O₂ reacts with a methine bridge of the haem, and that the bridge specificity is regulated by hindering amino acid residues that determine the location of the bound O₂. The residue phenylalanine-CD1 appears to be further away from the haem plane or in a markedly more flexible position in leghaemoglobins than in mammalian globins. The haem-bound oxygen atom B, in Fe–O(A)–O(B), seems to be free to rotate in all directions except that of the γ-bridge in Glycine and Phaseolus leghaemoglobins, but its position in Vicia and Pisum leghaemoglobin I might be restricted to the direction of the β-methine bridge.     

Structure and growth of infection threads in the legume symbiosis with Rhizobium leguminosarum

Specific antibodies and enzyme–gold probes were used to study the structure and development of infection threads in nodules induced by Rhizobium leguminosarum on the roots of Vicia, Pisum and Phaseolus. In Pisum nodules, the tubular infection thread wall contains polysaccharides antigenically similar to those of the cell wall, including cellulose, xyloglucan, methyl-esterified pectin and non-esterified pectin, but none of these wall components is present around the infection droplet structures from which bacteria are internalized by plant plasma membrane. As reported previously for pea nodules, the luminal matrix of infection threads and infection droplets contains a plant glycoprotein; this glycoprotein is also secreted by infected and uninfected cortical cells of a Vicia root at the earliest stages of nodule initiation. Synthesis of a transcellular infection thread apparently involves reorganized deposition of components normally targeted to the cell wall, and infection thread growth is orientated anticlinally through the outer cortex in the same plane observed for the deposition of new cell walls following mitosis. Both the development of infection threads in the outer cortex and the initiation of cell division in the inner cortex are preceded by a similar process of cell reactivation involving centralization of nuclei and the development of anticlinal transvacuolar strands. It is therefore suggested that the two Rhizobium-induced processes of infection thread growth and cortical cell division may both be consequences of a similar plant cell response in the inner and outer root cortex, respectively. Phaseolus nodules contained only short intracellular infection structures which terminated within individual cells and contained no luminal matrix material. The differences in infection thread structure between Pisum and Phaseolus nodules may reflect differences in ontogeny between “indeterminate” and “determinate” nodule meristems.     

A COMPARATIVE STUDY OF OVULE AND MEGAGAMETOPHYTE DEVELOPMENT IN FIELD-GROWN AND GREENHOUSE-GROWN PLANTS OF GLYCINE MAX AND PHASEOLUS AUREUS (PAPILIONACEAE)

A study of ovule and megagametophyte development in field- and greenhouse-grown plants of Glycine max (L.) Merrill and Phaseolus aureus Roxb. reveals several consistent features for both species. These features include: a multiple archesporium, enlargement of a primary sporogenous cell directly into a megasporocyte, production of unequal dyad cells, a functional chalazal megaspore, Polygonum-type development, and a hypostase. A filiform apparatus was not observed in either species. Several marked differences in development also occur. Phaseolus usually produces one sporogenous cell per ovule; Glycine produces 2–3 sporogenous cells per ovule. Meiosis II is synchronous in Phaseolus but nonsynchronous in Glycine. Linear tetrads are produced in Phaseolus, whereas linear and T-shaped tetrads are found in Glycine. Starch grains accumulate in the mature megagametophyte of Glycine but are absent at that stage in Phaseolus. The usefulness of the modified clearing fluid, benzyl benzoate-4½, for the study of ovule and megagametophyte development in Glycine max and Phaseolus aureus is here demonstrated. In addition, the study indicates for both species that megagametophyte development in plants grown under field conditions is markedly similar to development in plants grown in the more uniform conditions of the greenhouse. Accordingly, these findings suggest generally that embryological data collected from plants grown under greenhouse conditions will reflect those from plants found in nature.     

Some Effects of Sucrose Concentration on Primordium Development in Excised Primary Roots

The effects of varying the amount of sucrose used to supplementthe culture medium maintaining the growth of excised roots ofPisum sativum L., Vicia faba L., Zea mays L. and Phaseolus vulgarisL., on the rates of primordium initiation and subsequent emergenceas lateral roots and on the duration of the interval betweenprimordium inception and emergence as a secondary root throughthe tissues of the primary have been investigated. Variation in the exogenous concentration of sucrose from 0.5to 8 per cent had little effect on the rate of primordium inceptionin Pisum and Vicia and the rates never reached the values obtainedfor the roots of the corresponding intact plants. Moreover,over the 6 day culture period lateral root emergence did notoccur in any of the excised roots of these two species. In contrast,each of the aspects of primordium development examined in theexcised roots of Zea and Phaseolus was markedly affected bythe amount of sucrose used to supplement the culture medium.In addition, in the presence of about 6 per cent sucrose, primordiumdevelopment in these cultured roots was very similar to thatin roots of the corresponding intact plants. The results indicate either that some factor necessary for primordiumdevelopment is present in adequate amounts in excised rootsof Zea and Phaseolus, but not in those of Pisum and Vicia, orthat the factors controlling such development are differentin the former and latter two species. Vicia faba L., Pisum sativum L., Zea mays L., Phaseolus vulgaris L., broad bean, garden pea, maize, dwarf bean, primordium development, sucrose concentration, cultured roots     

The comparison of seed proteins of several representatives of the genusPisum with respect to their relationships An immunological comparison

Using immunochemical methods a comparison was made of a complex of water soluble (albumins) and salt soluble (globulins) seed proteins, especially vicilin and legumin, in selected species of the genusPisum, to determine the degree of their taxonomic relationship. Within the genusPisum the interspecific differences betweenP. abyssinicum, P. cinereum,P. elatius, P. fulvum, P. sativum, andP. syriacum are much smaller, and thus the taxonomic distances are shorter than is the casee.g. in the genusPhaseolus. In spite of this fact one may state thatPisum sativum, P. elatius andP. syriacum constitute a definite group, whereasP. abys-sinicum, P. cinereum and P.fulvum have longer taxonomic distances.     

Mild temperature “stress” and callose synthesis

Seedlings of Zea mays L., Sorghum vulgare, Pisum sativum L., Phaseolus aureus, Glycine max L. and Lycopersicum esculentum were grown at 20°C and at 26°C. The seedlings were fixed in glutaraldehyde and sections were examined for aniline-blue-induced fluorescence, which is supposedly indicative of -1,3-glucans or callose. There was much more aniline-blue fluorescence in Zea, Glycine and Phaseolus seedlings grown at 20°C compared with 26°C whereas Pisum and Lycopersicum seedlings grown at 26°C showed more fluorescence than those grown at 20°C. In Zea, large deposits of fluorescent material were particularly noticeable in the walls of elongating cells around the shoot apex and in root-cap cells, and appeared to be closely associated with a few of the pitfields. The remaining pitfields showed the normal, low level of aniline-blue fluorescence.     

Cellular pathway of photosynthate transport in coats of developing seed of Vicia faba L. and Phaseolus vulgaris L. I. Extent of transport through the coat symplast

The extent of post-phloem solute transport through the coatsymplasts of developing seeds of Vicia faba L. and Phaseolusvulgaris L. was evaluated. For Vicia seed coats, the membrane-impermeantfluorochrome, CF, moved radially from the chalazal vein to reachthe chlorenchyma and thin-walled parenchyma transfer cell layers.Thereafter, the fluorochrome moved laterally in these two celllayers around the entire circumference of the seed coat. Transferof CF from the chalazal vein was inhibited by plasmolysis ofattached ‘empty’ seed coats. In contrast, the spreadof phloem imported CF was restricted to the ground parenchymaof Phaseolus seed coats. Fluorochrome loaded into the outermostground parenchyma cell layer was rendered immobile followingplasmolysis of excised seed-coat halves. Phloem-imported [¹⁴C]sucroseand the slowly membrane permeable sugar, L-[¹⁴C]glucose, werepartitioned identically between the vascular and non-vascularregions of intact Vicia seed coats. For ¹⁴C-photosynthates,these partitioning patterns in attached ‘empty’Vicia seed coats were unaffected by PCMBS, but inhibited byplasmolysis. Tissue autoradiographs of intact Phaseolus seedcoats demonstrated that a pulse of ¹⁴C-photosynthate moved fromthe veins to the grounds tissues. In excised Vicia seed coats,preloaded with ¹⁴C-photosynthates, the cellular distributionof residual ¹⁴C-label was unaffected by PCMBS. In contrast,PCMBS caused the ¹⁴C-photosynthate levels to be elevated inthe veins and ground parenchyma relative to the branch parenchymaof Phaseolusseed coat halves. Based on the above findings, itis concluded that the phloem of Vicia seed coats is interconnectedto two major symplastic domains; one comprises the chlorenchyma,the other the thin-walled parenchyma plus thin-walled parenchymatransfer cells. For Phaseolusseed coats, the phloem forms amajor symplastic domain with the ground parenchyma. Key words: Phaseolus vulgaris L, phloem unloading, photosynthate transport, seed coat, symplast, Vicia faba L     

Measurements of genetic variability for symbiotic dinitrogen fixation in fieldgrwon fababean (Vicia faba L.) using a low level15N-tracer technique

Before starting a breeding program aimed at improving the nitrogen nutrition ofVicia faba, the authors tried an alternative technique to the acetylene reduction assay, to measure some genetic variability in the plant material. The quantity of dinitrogen fixed by several cultivars ofVicia faba was estimated using a low enrichment¹⁵N tracer method and high precision¹⁵N mass spectrometry. The fababeans were cultivated for two years in two different soils. The percentage of fixed dinitrogen in the seed varied between genotypes from 40 to 83% of the total nitrogen and was positively correlated with the total seed nitrogen (r=0.64 to 0.86). A highly significant positive correlation was also found between the total seed nitrogen and the quantity of fixed dinitrogen in the seed (r=0.95 to 0.99). The technique used to measure dinitrogen fixation proved to be useful and reliable enough to discriminate between various genotypes, grown over a period of two years in two different soils. However, several non-fixing control plants showed significant differences in their¹⁵N enrichment and the problem of choosing a good reference plant was raised and discussed.     

A QUANTITATIVE ANALYSIS OF FEMALE GAMETOPHYTE DEVELOPMENT IN FIELD- AND GREENHOUSE-GROWN PLANTS OF GLYCINE MAX AND PHASEOLUS AUREUS (PAPILIONACEAE)

A statistical analysis of size for the megasporocyte, functional megaspore, and 2-, 4-, 8-nucleate, and mature female gametophytes for Glycine max and Phaseolus aureus grown in the field and greenhouse was accomplished from measurements of the length, width, and length and width intercepts for each stage. The greatest increase in mean length for Phaseolus in the greenhouse and Glycine in the field takes place between the 2- and 4-nucleate stages. In alternate environments, the two genera show the greatest increase between the functional megaspore and 2-nucleate stage. Greater similarity between the genera than shown by each genus in the two environments was also found for other features, viz., the largest mean length and width attained by each stage, the least increase in mean length and width and the overlap in confidence intervals for length and length intercepts between successive stages, and changes in the width intercept as a percent of total width as the ovule becomes campylotropous. T tests at the 0.05 level reveal significant differences between greenhouse and field plants in Phaseolus for length of the megasporocyte and 8-nucleate stage and length and length intercept of the 2- and 4-nucleate stages. In Glycine, differences appear for the length and both intercepts of the megasporocyte and the length intercepts of the mature stage. The similarity of Glycine in one environment to Phaseolus in the other coupled with the differences of statistical significance for each genus in the two conditions suggest that environment does have a pronounced effect on female gametophyte development. Statements to the contrary in previous reports to include one on these genera are correct for the qualitative aspects of development investigated, but they cannot be extended in all respects to the quantitative analysis of growth reported here. The differences recorded suggest caution in the choice of material for comparative studies. For embryological data, especially those of a quantitative nature, to be fully useful in taxonomic assessments, material for all taxa should be collected from natural habitats.     

Development of Lateral Root Primordia in Vicia faba, Pisum sativum, Zea mays andPhaseolus vulgaris: Rates of Primordium Formation and Cell Doubling Times

Lateral root primordium development has been examined in primaryroots of Vicia faba L., Pisum sativum L., Zea mays L. and Phaseolusvulgaris L. Following their initiation from an estimated minimumnumber of 77–162, 20–57, 17 and 12 cells respectivelyin Vicia, Phaseolus, Pisum and Zea, the primordia rapidly increasedin cell number to emerge as secondary roots about 2.8–3.6days later depending on the species being examined. Cell doublingtimes were estimated directly from cell numbers at differenttimes following primordium inception and were found to increasewith increase in primordium size in each of the species investigated. The number of primordia formed per cm of root growth per daywas greatest in Zea and least in Pisum. A comparison of thedata obtained for Vicia with that in the literature led to theconclusion that although the number of primordia produced percm of root growth was independent of the rate of primary elongation,the number produced per day increased in a linear fashion withincrease in the rate at which the primary lengthened. Vicia faba L, Pisum sativum L, Zea mays L, Phaseolus vulgaris L, broad bean, garden pea, maize, dwarf bean, root primordia, cell division, cell doubling time     

Is nitrogen accumulation in grain legumes responsive to growth or ontogeny?

Nitrogen (N) accumulation in legumes is one of the main determinants of crop yield. Although N accumulation from symbiotic nitrogen fixation or N absorption from the soil has been widely investigated, there is no clear consensus on timing of the beginning of N accumulation and the termination of N accumulation and the physiological events that may be associated with these two events. The analyses conducted in this study aimed at identifying the determinant of N accumulation in two grain legume species. Nitrogen accumulation dynamics and mass accumulation and development stages were recorded in the field for several genotypes of common bean (Phaseolus vulgaris) and faba bean (Vicia faba) under different growing conditions. This study showed that during the vegetative stages, N accumulation rate was correlated with mass accumulation rate. However, the maximum accumulation of N did not correspond to the time of the maximum mass accumulation. In fact, for both species, N accumulation was found to persist in seed growth. This challenges a common hypothesis that seed growth causes a decrease in N accumulation because of a shift of the photosynthate supply to support the seed growth. Even more surprising was the shift of the active accumulation of N in faba bean to late in the growing season as compared with common bean. N accumulation by faba bean only was initiated at high rates very late in vegetative growth and persisted at high rates well into seed fill.     

Interspecific hybridization in the grain legumes—a review

The current position regarding reports in the literature on interspecific hybridization in the pulses is confused. Two major reasons can he advanced in explanation. The first is that frequently conspecific wild and cultivated forms have received different binomials and crosses between such are frequently regarded as interspecific hybrids. The second arises from mistaken interpretation of results of attempted interspecific hybridization when progeny strongly resembling the maternal parent are produced. It is suggested that these progeny might have been produced by failures in emasculation or rare accidental apomixis. All cultigens of the generaArachis, Cajanus, Cicer, Phaseolus andPisum are able to some extent to produce viable true interspecific hybrids as are the Asiatic forms ofVigna. True interspecific hybrids have not been produced withVicia faba nor withVigna unguiculata the cowpea.     

Dispersal mode constrains allocation of carbon and mineral nutrients in seeds of forest and savanna trees

• Species vary in seed size and content of stored reserves, which can be related to dispersal strategies and type of habitat in which they are found. We compare seed carbon and nutrient reserves of anemochorous and zoochorous trees from the Cerrado of central Brazil.
• We measured seed dry mass, lipids, non‐structural carbohydrates (starch and total soluble sugars), carbon and mineral nutrients in ten forest and 13 savanna species, each classified as having wind‐ or animal‐dispersed seeds. We used phylogenetically independent contrasts to test for correlations among these traits.
• Seeds of anemochorous species were lighter, with higher concentrations of C, N, P, Ca and Mg. Lipids were the dominant carbon reserve for most anemochorous species, underpinning the importance of allocation to compact carbon reserves. Starch, lipids or soluble sugars were the major carbon reserve in zoochorous seeds. Savanna and forest species did not differ in seed mass or in total carbon reserves. However, seeds of forest species had higher concentrations of starch than seeds of savanna species. Lipid and starch negatively correlated across species, suggesting a trade‐off between starch and lipids as major seed carbon reserves. Calcium was positively correlated with Mn and B, while Mg was positively correlated with C, N, P, K, S, Zn and B. Potassium, S and Cl were positively correlated, while P was positively correlated with Mg and Zn.
• Dispersal mode rather than vegetation type constrained seed mass and seed storage allocation patterns in forest and savanna trees. We provide evidence that similar mechanisms are involved in seed storage of carbon and mineral nutrients across species.

     

The cellular pathway of sucrose transport in developing cotyledons ofVicia faba L. andPhaseolus vulgaris L.: a physiological assessment

The cellular pathway of sugar uptake in developing cotyledons of Vicia faba L. and Phaseolus vulgaris L. seed was evaluated using a physiological approach. The cotyledon interface with the seed coat is characterised by a specialised dermal cell complex. In the case of Vicia faba cotyledons, the epidermal component of the dermal cell complex is composed of transfer cells. Sucrose is the major sugar presented to the outer surface of both cotyledons and it is taken up from the apoplasm unaltered. Estimated sucrose concentrations within the apparent free space of Vicia and Phaseolus cotyledons were 105 and 113 mM respectively. Rates of in-vitro uptake of [¹⁴C]sucrose by cotyledon segments or by whole cotyledons following physical removal or porter inactivation of the outer cells demonstrated that, for both Vicia and Phaseolus cotyledons, the dermal cell complexes are the most intense sites of sucrose uptake. Accumulation of [¹⁴C]sucrose in the storage parenchyma of whole cotyledons was directly affected by experimental manipulation of uptake by the outer cell layers and plasmolytic disruption of the interconnecting plasmodesmata. These findings indicated that sucrose accumulated by the dermal cell complexes is transported symplasmically to the storage parenchyma. Overall, it is concluded that the dermal cell complexes of the developing legume embryo, irrespective of the presence or absence of wall ingrowths, are the major sites for the uptake of sucrose released from the maternal tissues to the seed apoplasm. Thereafter, the accumulated sucrose is transported radially inward through the symplast to the storage parenchyma.Abbreviations AFS apparent free space - CF 5-(6)-carboxyfluorescein - CFDA 5-(6)-carboxyfluorescein diacetate - Mes 2-(N-morpholino)ethanesulfonic acid - PCMBS p-chloromercuribenzenesulfonic acid - SRG sulphorhodamine G The investigation was supported by funds from the Research Management Committee, The University of Newcastle and the Australian Research Council. One of us, R. McDonald, gratefully acknowledges the support of an Australian Postgraduate Research Award. We are grateful to Stella Savoury for preparing the photomicrographs.     

Nitrogen Nutrition of Nodules in Relation to ;N-Hunger' in Cowpea (Vigna unguiculata L. Walp)

Early growth, nodule development, and nitrogen fixation by two cultivars of cowpea (Vigna unguiculata L. Walp), one large-seeded (Vita 3; 146.0 ± 0.9 milligrams seed dry weight, 4.1 ± 0.2 milligrams seed N), the other small-seeded (Caloona; 57.5 ± 2.5 milligrams seed dry weight, 1.8 ± 0.1 milligrams seed N), were compared under conditions of sand culture with nutrient solution free of combined N. The seed stocks used had been obtained from plants uniformly labeled with ¹⁵N, thus enabling changes with time in distribution of cotyledon and fixed N among plant parts to be measured by isotope dilution. Caloona, but not Vita 3, showed physiological symptoms of `N hunger,' i.e. transient loss of chlorophyll (visible yellowing) and N from the first-formed unifoliolate leaves at or around the onset of symbiotic functioning and N<sub>2</sub> fixation. The smaller-seeded Caloona showed higher early nitrogenase activity than the larger-seeded Vita 3 and by 28 days had fixed 6.6 milligrams of N per milligram of seed N [mg N · (mg seed N)<sup>−1</sup>] versus only 3.5 mg N · (mg seed N)<sup>−1</sup> in Vita 3. Both cultivars lost around 30% of their initial seed N at germination, mostly as fallen cotyledons. Abscised cotyledons of Caloona contained 1.21 ± 0.17% N; those of Vita 3 contained 2.61 ± 0.37% N. When compared on the basis of cotyledon N available for seedling growth, Caloona was shown to have fixed 10.6 mg N · (mg seed N)<sup>−1</sup> and Vita 3 only 5.3 mg N · (mg seed N)<sup>−1</sup>. Most of the cotyledon N withdrawn from the unifoliolate leaf pair of Caloona during `N-hunger' was committed to early nodule growth and, in total, 20 to 25% of the cotyledon N resource of this cultivar was ultimately invested in establishment of symbiosis compared with only 7% in Vita 3.     

Turgor-dependent unloading of photosynthates from coats of developing seed of Phaseolus vulgaris and Vicia faba. Turgor homeostasis and set points

Key physiological characteristics of turgor-dependent efflux of photosynthates were examined using excised coats and cotyledons of developing Phaseolus vulgaris (cv. Redland Poineer) and Vicia faba (cv. Coles Prolific) seed during the linear phase of seed fill. Exposure to solutions of high osmotic potential inhibited net uptake of [¹⁴C]sucrose by cotyledons at developmental stages less than 60% of their final dry weight. The effect could not be fully reversed by transferring cotyledons to solutions set at lower osmotic potentials. The inhibition became apparent at osmotic potentials that were higher than those that caused stimulation of efflux from seed coats. Net [¹⁴C]sucrose uptake by cotyledons at more advanced stages of development was unaffected by external osmotic potential. Specified tissue layers were removed from seed coats by pretreatment with pectinase. Efflux studies with the pectinase-modified coats of Phaseolus and Vicia seed demonstrated that the cellular site of turgordependent efflux was the ground parenchyma and thin-wall parenchyma transfer cells, respectively. Coats subjected to long-term (hours) incubations, under hypo-osmotic conditions, exhibited the capacity for turgor regulation. This was mediated by turgor-dependent efflux of solutes. The solutes exchanged were of nutritional significance to the developing embryo. The relationship between efflux and coat turgor was characterised by a turgor-independent phase at low turgors. Once turgor exceeded a minimal value (set point), efflux increased in proportion to the magnitude of the turgor deviation (error signal) from the set point. For coats of sink-limited seed of Vicia and Phaseolus, efflux exhibited apparent saturation at turgors above 0.25 and 0.5 MPa respectively. The putative turgor set point and slope of the turgor-dependent component of efflux varied with seed development, the prevailing source/sink ratio and genetic differences in seed growth rate. The nature of these kinetic variations was compatible with the competitive ability of the seed. A turgor homeostat model is proposed that incorporates the observed functional attributes of turgor-dependent efflux. Operationally, the model provides a mechanistic basis for the integration of assimilate demand by the cotyledons with assimilate import into and unloading from the seed coat.     

Evolutionary analysis of the APA genes in the <Emphasis Type="Italic">Phaseolus</Emphasis> genus: wild and cultivated bean species as sources of lectin-related resistance factors?

The APA (Arcelin/Phytohemagglutinin/α-Amylase inhibitor) gene family is composed of various members, present in Phaseolus species and coding for lectin and lectin-related seed proteins having the double role of storage and defense proteins. Here members of the APA family have been identified by immunological, functional, and molecular analyses and representative genes were sequenced in nine wild species of Phaseolus. All taxa possessed at least one member of the true lectin gene. No arcelin type sequences have been isolated from the species examined. Among the wild species studied, only P. costaricensis contained an α-amylase inhibitor (α-AI). In addition P. augusti, P. maculatus, P. microcarpus, and P. oligospermus showed the presence of the lectin-related α-amylase inhibitor-like (AIL) genes and α-AI activity. Data from Southern blot analysis indicated the presence of only one lectin gene in P. parvulus and P. filiformis, while an extensive gene duplication of the APA locus was found in the other Phaseolus species. Phylogenetic analysis carried out on the nucleotide sequences showed the existence of two main clusters and clearly indicated that lectin-related genes originated from a paralogous duplication event preceding the development of the ancestor to the Phaseolus genus. The finding of detectable α-AI activity in species containing AIL genes suggests that exploiting APA genes variability in the Phaseolus genus may represent a valuable tool to find new members that may have acquired insecticidal activities. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Genetic diversity of SIRE-1 retroelements in annual and perennial glycine species revealed using SSAP

Sequence Specific Amplification Polymorphisms (SSAP) were used to measure the distribution and structure of SIRE-1 retroelement populations in annual and perennial Glycine species. For SSAP analysis, primers corresponding to a region immediately upstream of the 3’LTR of the soybean retroelement SIRE-1 were chosen. Analysis reveals that SIRE-1 is present throughout the Glycine genus and shows that the annual species have similar SIRE-1 populations whilst the perennial species have much more distinct and diverse populations. The high number of species-specific subgroups suggest that SIRE-1 has been active and evolving independently in each species during the course of Glycine evolution.     

Environmental effects on photosynthesis, nitrogen-use efficiency, and metabolite pools in leaves of sun and shade plants

Effects of varying light intensity and nitrogen nutrition on photosynthetic physiology and biochemistry were examined in the sun plant Phaseolus vulgaris (common bean) and in the shade plant Alocasia macrorrhiza (Australian rainforest floor species). In both Phaseolus and Alocasia, the differing growth regimes produced large changes in photosynthetic capacity and composition of the photosynthetic apparatus. CO₂-saturated rates of photosynthesis were linearly related to leaf nitrogen (N) content in both species but photosynthesis per unit leaf N was markedly higher for Phaseolus than for Alocasia. Photosynthetic capacity was also higher in Phaseolus per unit ribulose 1,5-bisphosphate (RuBP) carboxylase (RuBPCase) protein. The leaf content of RuBPCase was linearly dependent on leaf N content in the two species. However, the proportion of leaf N which was RuBPCase was greater in Phaseolus than in Alocasia and was more sensitive to growth conditions, ranging from 6% of leaf N at low light to 20% at high light. In Alocasia, this range was much less, 6 to 11%. However, chlorophyll content was much more sensitive to light intensity in Alocasia. Thus, the RuBPCase/chlorophyll ratio was quite responsive to N availability and light intensity in both species (but for different reasons), ranging from 6 grams per gram for Phaseolus and 2 grams per gram for Alocasia at high leaf N and 1.5 gram per gram for Phaseolus and 0.5 gram per gram for Alocasia at low leaf N. These large changes in the proportions of components of the photosynthetic apparatus had marked effects on the sensitivity of these species to photoinhibition. These environmental effects also caused changes in the absolute levels of metabolites of the photosynthetic carbon reduction cycle. Concentrations of RuBP and P-glycerate were approximately 2-fold higher in high light-grown than low light-grown Phaseolus and Alocasia when expressed on a leaf area basis. However, if metabolite pool sizes are expressed on the basis of the RuBPCase catalytic site concentration, then they were little affected by the marked changes in leaf makeup. There appears to be fundamental differences between these species in the mechanism of sun-shade adaptation and N partitioning in the photosynthetic apparatus that result in significant differences in the N-use efficiency of photosynthesis between Phaseolus and Alocasia but similar RuBPCase:substrate:product ratios despite these differences.