Origin and differentiation of Aegilops triuncialis L. as determined by esterase isozyme analysis

Summary Banding patterns of esterase isozymes in Aegilops triuncialis (2n = 28, genome formula C^uC^uCC) and its putative parental species, Ae. umbellulata (2n = 14, C^uC^u) and Ae. caudata (2n = 14, CC), were studied by the gel isoelectric focusing method using pH 6–8 carrier ampholite. Zymogram phenotypes of both parents were quite uniform. Seven zymogram phenotypes (designated as phenotypes 1 to 7) were found among the 260 strains of Ae. triuncialis examined. Of these phenotypes, phenotype 1 was identical to the zymogram phenotype produced by the ancestral species, Ae. umbellulata, and bands considered to have been derived from Ae. caudata were absent in this phenotype. Phenotype 3 had all bands of both parents. The other phenotypes differed greatly from phenotype 3. Therefore, phenotype 3 was considered to be most primitive of the 7 types, and the Ae. triuncialis strains which showed phenotype 3 to be the most primitive of the strains examined. If Ae. triuncialis originated as a hybrid between Ae. umbellulata and Ae. caudata, the zymogram phenotype must have been phenotype 3, in which the isozymes of both parental species are present. Whether the phenotypes other than type 3 were due to introgressive hybridization could not be verified, but they were considered in this article to be a consequence of a rearrangement of chromosomes.Contribution from the Laboratory of Genetics, Faculty of Agriculture, Kyoto University No. 432     

Meiotic restriction in emmer wheat is controlled by one or more nuclear genes that continue to function in derived lines

Highly fertile F₁ hybrids were made between Triticum turgidum L. ssp. turgidum (2n = 28, AABB) and Aegilops tauschii Coss. (2n = 14, DD) without embryo rescue and hormone treatment. The F₁ plants had an average seed set of 25%. Approximately 96% of the F₂ seeds were able to germinate normally and about 67% of the F₂ plants were spontaneous amphidiploid (2n = 42, AABBDD). Cytological analysis of male gametogenesis of the F₁ plants showed that meiotic restitution is responsible for the high fertility. A mitosis-like meiosis led to meiotic restitution at either of the two meiotic divisions resulting in unreduced gametes. Test crosses of the T. t. turgidum–Ae. tauschii amphidiploid with Ae. variabilis and rye suggested that the mitosis-like meiosis is controlled by one or more nuclear genes that continue to function in derived lines. This discovery indicates a potential application of such genes in producing double haploids.     

Analysis of the resistance of Aegilops squarrosa to the wheatgrass mildew fungus by using the gene-for-gene reationship

Summary Pm10 and Pm15, resistance genes to Erysiphe graminis f. sp. agropyri, are located on the D genome of common wheat. It was determined whether or not they were carried by existing lines of the D genome donor, Aegilops squarrosa, using the gene-for-gene relationship. Two lines of Ae. squarrosa tested (one was var. meyeri and the other was var. anathera) were susceptible to culture Tk-1 of E. graminis f. sp. tritici and were highly resistant to culture Ak-1 of E. graminis f. sp. agropyri. The two lines were inoculated with an F₁ population derived from the cross Ak-1 × Tk-1. Comparative analyses of the segregation patterns revealed that Ppm10 and Ppm15, avirulence genes corresponding to Pm10 and Pm15, respectively, are involved in the avirulence of Ak-1 on var. meyeri and var. anathera, respectively. According to the gene-for-gene relationship, var. meyeri and var. anathera were inferred to carry Pm10 and Pm15, respectively. Analysis with a synthetic hexaploid confirmed the inference.     

Plant regeneration from embryogenic cultures initiated from mature loblolly pine zygotic embryos

Summary Mature zygotic embryos of eight (open-pollinated) families of loblolly pine (Pinus taeda L.) were cultured on eight different basal salt formulations, each supplemented with 36.2 μM 2,4-dichlorophenoxyacetic acid, 17.8 μM 6-benzyladenine, 18.6 μM kinetin, 500 mg l⁻¹ casein hydrolysate, and 500 mg l⁻¹ l-glutamine for 9 wk; embryogenic tissue was formed on cotyledons, hypocotyls, and radieles of mature zygotic embryos. Callus was subcultured on the callus proliferation medium, the same as the induction medium but with one-fifth concentration of auxin and cytokinin for 9 wk. On this medium a white to translucent, glossy, mucilaginous embryogenic callus containing embryogenic suspensor masses (ESMs) was obtained. The highest frequency of explants forming embryogenic tissue, 17%, occurred on a modified Murashige and Skoog salts basal medium containing the concentration of KNO₃, Ca(NO₃)₂·4H₂O, NH₄NO₃, KCl, ZnSO₄·7H₂O, and MnSO₄·H₂O, 720, 1900, 400, 250, 25.8, and 25.35 mg l⁻¹, respectively. Embryogenic suspension cultures were established by culturing embryogenic callus in liquid callus proliferation medium. Liquid cultures containing ESMs were transferred to medium containing abscisic acid, polyethylene glycols, and activated charcoal for stimulating the production of cotyledonary somatic embryos. Mature somatic embryos germinated for 4–12 wk on medium containing indole-butyric acid, gibberellic acid, 6-benzyladenine, activated charcoal, and reduced sucrose concentration (15 g l⁻¹). Two hundred and ninety-one regenerated plantlets were transferred to a perlite:peatmoss:vermiculite (1∶1∶1) mixture, then the plants were transplanted to soil in the earth, and 73 plantlets survived in the field.     

Standard karyotypes ofAegilops uniaristata,Ae. mutica,Ae. comosa subspeciescomosa andheldreichii (Poaceae)

C-banding patterns and polymorphisms were analyzed in several accessions of the diploidAegilops speciesAe. uniaristata, Ae. mutica, andAe. comosa subsp.comosa and subsp.heldreichii, and standard karyotypes of these species were established. Variation in C-band size and location was observed between different accessions, but did not prevent chromosome identification. One accession ofAe. uniaristata was homozygous for whole-arm translocations involving chromosomes 1N and 5N. The homoeologous relationships of these chromosomes were established by comparison of chromosome morphologies and C-banding patterns to other diploidAegilops species with known chromosome homoeology. In addition, in situ hybridization analysis with a 5S rDNA probe was used to identify homoeologous groups 1 and 5 chromosomes. The present analysis permitted the assignment of allAe. mutica, comosa subsp.comosa, andAe. comosa subsp.heldreichii chromosomes, and three of the sevenAe. uniaristata chromosomes according to their homoeologous groups. The data presented will be useful analyzing genome differentiation in polyploidAegilops species.     

Effect of solarization of soil within plastic bags on root rot of gerbera (Gerbera jamesonii L.)

Solarization of soil, (potting mix = coarse sand:Eucalyptus marginata fines = 1∶1) infested with 3 fungi pathogenic to gerbera (Phytophthora cryptogea, Fusarium oxysporum andRhizoctonia solani), for 3 to 4 weeks within transparent polyethylene bags controlled root rot of gerbera. Solarization for 2 weeks however, was less effective. All plants grown in the infested potting mix which had been kept in shade for 2, 3 or 4 weeks were severely attacked. Solarization of soil within plastic bags for 4 weeks also increased availability of nutrients such as NH₄ ⁺-N, PO₄ ⁻ and K⁺ in comparison to bagged soil kept in the shade for the same period.     

Relationships between the chromosomes of Aegilops umbellulata and wheat

 A comparative genetic map of Aegilops umbellulata with wheat was constructed using RFLP probes that detect homoeoloci previously mapped in hexaploid bread wheat. All seven Ae. umbellulata chromosomes display one or more rearrangements relative to wheat. These structural changes are consistent with the sub-terminal morphology of chromosomes 2 U, 3 U, 6 U and 7 U. Comparison of the chromosomal locations assigned by mapping and those obtained by hybridization to wheat/Ae. umbellulata single chromosome addition lines verified the composition of the added Ae. umbellulata chromosomes and indicated that no further cytological rearrangements had taken place during the production of the alien-wheat aneuploid lines. Relationships between Ae. umbellulata and wheat chromosomes were confirmed, based on homoeology of the centromeric regions, for 1 U, 2 U, 3 U, 5 U and 7 U. However, homoeology of the centromeric regions of 4 U with wheat group-6 chromosomes and of 6 U with wheat group-4 chromosomes was also confirmed, suggesting that a re-naming of these chromosomes may be pertinent. The consequences of the rearrangements of the Ae. umbellulata genome relative to wheat for gene introgression are discussed. Received: 10 July 1997 / Accepted: 19 September 1997     

Electrophoretic variation in esterases and peroxidases of native greek diploidAegilops species(Ae. caudata andAe. comosa,Poaceae)

The esterase and peroxidase patterns in five varieties ofAegilops caudata (genome type C) andAe. comosa (genome type M) were studied in order to elucidate the phylogenetic relationships within and between the two groups. The electrostarch gel electrophoresis technique was applied to extracts of shoot and root of 4-day-old seedlings, and the electropherograms were evaluated by gel densitometer traces. Inspite of considerable isozyme polymorphism, closer relationships in the banding patterns were found between different varieties of a single species than between varieties of the two different species. Esterase and peroxidase patterns of the twoAe. caudata varieties (caudata andpolyathera) are very similar and prove their close phylogenetic relationship. The isozyme affinities withinAe. comosa varieties are illustrated by the seriessubventricosa—biaristata—thessalica. The latter endemic variety has quite a number of characteristic bands and is relatively isolated. Altogether, the electrophoretic data agree well with morphological and cytological similarities (Zhukovsky 1928,Eig 1929,Karataglis 1973, 1975b).     

A chromosome-specific sequence common to the B genome of polyploid wheat and Aegilops searsii

A low-copy, non-coding chromosome-specific DNA sequence, isolated from common wheat, was physically mapped to the distal 19% region of the long arm of chromosome 3B (3BL) of common wheat. This sequence, designated WPG118, was then characterized by Southern hybridization, PCR amplification and sequence comparison using a large collection of polyploid wheats and diploid Triticum and Aegilops species. The data show that the sequence exists in all polyploid wheats containing the B genome and absent from those containing the G genome. At the diploid level, it exists only in Ae. searsii, a diploid species of section Sitopsis, and not in other diploids including Ae. speltoides, the closest extant relative to the donor of the B genome of polyploid wheat. This finding may support the hypothesis that the B-genome of polyploid wheat is of a polyphyletic origin, i.e. it is a recombined genome derived from two or more diploid Aegilops species.     

Comparative genetic analysis of the Aegilops longissima and Ae. sharonensis genomes with common wheat

Aegilops longissima Schw. et Musch. (2n= 2x=14, S^lS^l) and Aegilops sharonensis Eig. (2n=2x=14, S^lS^l) are diploid species belonging to the section Sitopsis in the tribe Triticeae and potential donors of useful genes for wheat breeding. A comparative genetic map was constructed of the Ae. longissima genome, using RFLP probes with known location in wheat. A high degree of conserved colinearity was observed between the wild diploid and basic wheat genome, represented by the D genome of cultivated wheat. Chromosomes 1S^l, 2S^l, 3S^l, 5S^l and 6S^l are colinear with wheat chromosomes 1D, 2D, 3D, 5D and 6D, respectively. The analysis confirmed that chromosomes 4S^l and 7S^l are translocated relative to wheat. The short arms and major part of the long arms are homoeologous to most of wheat chromosomes 4D and 7D respectively, but the region corresponding to the distal segment of 7D was translocated from 7S^lL to the distal region of 4S^lL. The map and RFLP markers were then used to analyse the genomes and added chromosomes in a set of ’Chinese Spring’ (CS)/Ae. longissima chromosome additions. The study confirmed the availability of disomic CS/Ae. longissima addition lines for chromosomes 1S^l, 2S^l, 3S^l, 4S^l and 5S^l. An as yet unpublished set of Ae. sharonensis chromosome addition lines were also available for analysis. Due to the gametocidal nature of Ae. sharonensis chromosomes 2S^l and 4S^l, additions 1S^l, 3S^l, 5S^l, 6S^l and 7S^l were produced in a (4D)4S^l background, and 2S^l and 4S^l in a euploid wheat background. The analysis also confirmed that the 4/7 translocation found in Ae. longissima was not present in Ae. sharonensis although the two wild relatives of wheat are considered to be closely related. The phenotypes of the Ae. sharonensis addition lines are described in an Appendix. Received: 28 September 2000 / Accepted: 19 January 2001     

D genome doners for Aegilops cylindrica (CCDD) and Triticum aestivum (AABBDD) deduced from esterase isozyme analysis

Summary Putative D genome donors for Aegilops cylindrica (2n = 28, CCDD) and Triticum aestivum (2n = 42, AABBDD) were studied with the isoelectric focusing patterns of esterase isozymes. 103 strains of Ae. cylindrica were uniform in their isozyme pattern. 30 strains of the putative parent, Ae. caudata, showed no zymogram variation, whereas the other parent, Ae. squarrosa, comprised 3 phenotypes. Natural Ae. cylindrica had an isozyme pattern which corresponded to a mixture of esterases from Ae. caudata and type 3 Ae. squarrosa. Therefore, it is concluded that the D genome donor of Ae. cylindrica is derived from type 3 Ae. squarrosa. These results suggest that Ae. cylindrica originated with a single amphiploidy event, and the C and D genomes have remained remarkably constant regarding esterase isozyme composition.On the other hand, T. aestivum comprised three zymogram phenotypes. These phenotypes contain bands which can be ascribed to the D genome of type 2 Ae. squarrosa. These results suggest that the D genome of Ae. cylindrica differs from that of T. aestivum. Evolution of the AB and D genomes of T. aestivum is indicated by the zymogram polymorphism. The origin of Ae. cylindrica is possibly more recent than that of T. aestivum.Contribution No. 433 of the Laboratory of Genetics, Faculty of Agriculture, Kyoto University     

Grain protein variability among species of Triticum and Aegilops: quantitative SDS-PAGE studies

Summary Total proteins were extracted from degermed seeds of various species of Triticum and Aegilops with solutions containing sodium dodecyl sulfate (SDS) and mercaptoethanol. The reduced, dissociated proteins were fractionated according to molecular weight (MW) by high-resolution polyacrylamide gel electrophoresis in buffers containing SDS (SDS-PAGE). Stained SDS-PAGE patterns were measured by densitometric scanning over a suitable range of optical density. The data were normalized to equivalent total areas for each of the densitometric scans by means of a computer program that also permitted the construction of patterns of hypothetical amphiploids by averaging patterns of two or three diploid species. The grain proteins of most species examined had distinctive qualitative and quantitative aspects that were characteristic of the species even though nearly every accession or cultivar of a species exhibited at least minor differences in pattern from other accessions or cultivars. The main protein components (probably prolamins) of Triticum monococcum ssp. monococcum, T. monococcum ssp. boeoticum, T. urartu, and Aegilops squarrosa had MW's in the range 29–36 X 10³ whereas the most important components of Ae. speltoides, Ae. longissima, and Ae. searsii had MW's in the range 37–55 × 10³. Changes in the quantitative expression of particular genes, especially those coding for storage protein components, may have been associated with speciation. The strong predominance of proteins with MW's in the range 29–36 × 10³ in some accessions of AB genome tetraploids, such as T. turgidum ssp. dicoccoides, may indicate contributions to the B genome of these tetraploids by T. monococcum ssp. boeoticum, T. urartu, or Ae. squarrosa.     

Statistical optimization of culture media for production of phycobiliprotein by Synechocystis sp. PCC 6701

Synechocystis sp. PCC 6701 has a brilliantly colored pigment, phycobiliprotein containing phycoerythrin. Culture medium was optimized by sequential designs in order to maximize phycobiliprotein production. The observed fresh weights after 6 days were 0.58 g/L in BG-11, 0.83 g/L in medium for Scenedesmus sp. and 0.03∼0.52 g/L in the other tested media. Medium for Scenedesmus sp. was selected to be optimized by fractional factorial design and central composite design since the medium maintained a more stable pH within a desirable range due to higher contents of phosphate. The fractional factorial design had seven factors with two levels: KNO₃, NaNO₃, NaH₂PO₄, Na₂HPO₄, Ca(NO₃)₂, FeEDTA, and MgSO₄. From the result of fractional factorial design, nitrate and phosphate were identified as significant factors. A central composite design was then applied with four variables at five levels each: nitrate, phosphate, pH, and light intensity. Parameters such as fresh weight and phycobiliprotein contents were used to determine the optimum value of the four variables. The proposed optimum media contains 0.88 g/L of nitrate, 0.32 g/L of phosphate under 25 μE·m⁻²·s⁻¹ of light intensity. The maximum phycobiliprotein contents have been increased over 400%, from 4.9 to 25.9 mg/L after optimization.     

Aspartate aminotransferase and alcohol dehydrogenase isoenzymes: Intraspecific differentiation inAegilops tauschii and the origin of the D genome polyploids in the wheat group

Polyacrylamide gel electrophoresis of aspartate aminotransferase (AAT, EC 2.6.1.1) and alcohol dehydrogenase (ADH, EC 1.1.1.1) isoenzymes reveals intraspecific differentiation ofAegilops tauschii Coss. (=Ae. squarrosa auct., non L.) into two groups of biotypes which essentially correspond to its two morphological subspecies, subsp.tauschii and subsp.strangulata (Eig)Tzvel. Subsp.tauschii which is characterized by a slower electromorph of AAT-B and a faster electromorph of ADH-A is identified as the contributor of its D genome to the tetraploidAe. cylindrica Host and the hexaploidAe. crassa Boiss. subsp.crassa. Subsp.strangulata, being distinguished by a faster electromorph of AAT-B and a slower electromorph of ADH-A, has contributed the D genome to the hexaploid bread wheats (Triticum aestivum L. emend.Thell.), the tetraploidsAe. crassa subsp.macrathera (Boiss.)Zhuk. andAe. ventricosa Tausch, and the hexaploidAe. juvenalis (Thell.)Eig. —Aegilops comosa Sibth. etSm. s. lat. is questioned as the contributor of the M genome toAe. crassa. Furthermore, the S genome diploidsAe. bicornis (Forsk.)Jaub. & Spach,Ae. longissima Schweinf. & Muschl. s. lat. andAe. searsii Feldman & Kislev are all considered unsuitable as the wheat B genome donors on the basis of the AAT isoenzyme data.     

Atrazine-resistant cytoplasmic male-sterile-nigra broccoli obtained by protoplast fusion between cytoplasmic male-sterile Brassica oleracea and atrazine-resistant Brassica campestris

Summary By using restriction endonuclease digestion patterns, the degree of intraspecific polymorphism of mitochondrial DNA in four diploid species of wheat and Aegilops, Ae. speltoides, Ae. longissima, Ae. squarrosa, and Triticum monococcum, was assessed. The outbreeding Ae. speltoides was found to possess the highest degree of variability, the mean number of nucleotide substitutions among conspecific individuals being 0.027 substitutions per nucleotide site. A very low degree of mtDNA variation was detected among Ae. longissima accessions, with most of the enzyme-probe combinations exhibiting uniform hybridization patterns. The mean number of substitutions among Ae. longissima individuals was 0.001 substitutions per nucleotide site. The domesticated diploid wheat T. monococcum var. monococcum and its conspecific variant T. monococcum var. boeoticum seem to lack mitochondrial DNA variability altogether. Thus, the restriction fragment pattern can be used as a characteristic identifier of the T. monococcum cytoplasmic genome. Similarly, Ae. squarrosa accessions were found to be genetically uniform. A higher degree of variation among accessions is observed when noncoding sequences are used as probes then when adjacent coding regions are used. Thus, while noncoding regions may contain regulatory functions, they are subject to less stringent functional constraints than protein-coding regions. Intraspecific variation in mitochondrial DNA correlates perfectly with the nuclear variability detected by using protein electrophoretic characters. This correlation indicates that both types of variation are selectively neutral and are affected only by the effective population size.     

Genome differentiation in Aegilops. 3. Evolution of the D-genome cluster

 Six polyploid Aegilops species containing the D genome were studied by C-banding and fluorescence in situ hybridization (FISH) using clones pTa71 (18S-5.8S-26S rDNA), pTa794 (5S rDNA), and pAs1 (non-coding repetitive DNA sequence) as probes. The C-banding and pAs1-FISH patterns of Ae. cylindrica chromosomes were identical to those of the parental species. However, inactivation of the NOR on chromosome 5D with a simultaneous decrease in the size of the pTa71-FISH site was observed. The N^v and D^v genomes of Ae. ventricosa were somewhat modified as compared with the N genome of Ae. uniaristata and the D genome of Ae. tauschii. Modifications included minor changes in the C-banding and pAs1-FISH patterns, complete deletion of the NOR on chromosome 5D^v, and the loss of several minor 18S-5.8S-26S rDNA loci on N^v genome chromosomes. According to C-banding and FISH analyses, the D^cr1 genome of Ae. crassa is more similar to the D^v genome of Ae. ventricosa than to the D genome of Ae. tauschii. Mapping of the 18S-5.8S-26S rDNA and 5S rDNA loci by multicolor FISH suggests that the second (X^cr) genome of tetraploid Ae. crassa is a derivative of the S genome (section Emarginata of the Sitopsis group). Both genomes of Ae. crassa were significantly modified as the result of chromosomal rearrangements and redistribution of highly repetitive DNA sequences. Hexaploid Ae. crassa and Ae. vavilovii arose from the hybridization of chromosomal type N of tetraploid Ae. crassa with Ae. tauschii and Ae. searsii, respectively. Chromosomal type T1 of tetraploid Ae. crassa and Ae. umbellulata were the ancestral forms of Ae. juvenalis. The high level of genome modification in Ae. juvenalis indicates that it is the oldest hexaploid species in this group. The occurrence of hexaploid Ae. crassa was accompanied by a species-specific translocation between chromosomes 4D^cr1 and 7X^cr. No chromosome changes relative to the parental species were detected in Ae. vavilovii, however, its intraspecific diversity was accompanied by a translocation between chromosomes 3X^cr and 3D^cr1. Received July 24, 2001 Accepted October 1, 2001     

NADP-dependent aromatic alcohol dehydrogenase in polyploid wheats and their diploid relatives. On the origin and phylogeny of polyploid wheats

Summary The three major isoenzymes of the NADP-dependent aromatic alcohol dehydrogenase (ADH-B), distinguished in polyploid wheats by means of polyacrylamide gel electrophoresis, are shown to be coded by homoeoalleles of the locus Adh-2 on short arms of chromosomes of the fifth homoeologous group. Essentially codominant expression of the Adh-2 homoeolleles of composite genomes was observed in young seedlings of hexaploid wheats (T. aestivum s.l.) and tetraploid wheats of the emmer group (T. turgidum s.l.), whereas only the isoenzyme characteristic of the A genome is present in the seedlings of the timopheevii-group tetraploids (T. timopheevii s.str. and T. araraticum).The slowest-moving B³ isoenzyme of polyploid wheats, coded by the homoeoallele of the B genome, is characteristic of the diploid species Aegilops speltoides S.l., including both its awned and awnless forms, but was not encountered in Ae. bicornis, Ae. sharonensis and Ae. longissima. The last two diploids, as well as Ae. tauschii, Ae. caudata, Triticum monococcum s.str., T. boeoticum s.l. (incl. T. thaoudar) and T. urartu all shared a common isoenzyme coinciding electrophoretically with the band B² controlled by the A and D genome homoeoalleles in polyploid wheats. Ae. bicomis is characterized by the slowest isoenzyme, B⁴, not found in wheats and in the other diploid Aegilops species studied.Two electrophoretic variants of ADH-B, B¹ and B², considered to be alloenzymes of the A genome homoeoallele, were observed in T. dicoccoides, T. dicoccon, T. turgidum. s.str. and T. spelta, whereas B² was characteristic of T. timopheevii s.l. and only B¹ was found in the remaining taxa of polyploid wheats. The isoenzyme B¹, not encountered among diploid species, is considered to be a mutational derivative which arose on the tetraploid level from its more ancestral form B² characteristic of diploid wheats.The implication of the ADH-B isoenzyme data to the problems of wheat phylogeny and gene evolution is discussed.     

Dinitrogen-fixation by three neotropical agroforestry tree species under semi-controlled field conditions

Cultivating dinitrogen-fixing legume trees with crops in agroforestry is a relatively common N management practice in the Neotropics. The objective of this study was to assess the N₂ fixation potential of three important Neotropical agroforestry tree species, Erythrina poeppigiana, Erythrina fusca, and Inga edulis, under semi-controlled field conditions. The study was conducted in the humid tropical climate of the Caribbean coastal plain of Costa Rica. In 2002, seedlings of I. edulis and Vochysia guatemalensis were planted in one-meter-deep open-ended plastic cylinders buried in soil within hedgerows of the same species. Overall tree spacing was 1 × 4 m to simulate a typical alley-cropping design. The ¹⁵N was applied as (NH₄)₂SO₄ at 10% ¹⁵N atom excess 15 days after planting at the rate of 20 kg [N] ha⁻¹. In 2003, seedlings of E. poeppigiana, E. fusca, and V. guatemalensis were planted in the same field using the existing cylinders. The ¹⁵N application was repeated at the rate of 20 kg [N] ha⁻¹ 15 days after planting and 10 kg [N] ha⁻¹ was added three months after planting. Trees were harvested 9 months after planting in both years. The ¹⁵N content of leaves, branches, stems, and roots was determined by mass spectrometry. The percentage of atmospheric N fixed out of total N (%N_f) was calculated based on ¹⁵N atom excess in leaves or total biomass. The difference between the two calculation methods was insignificant for all species. Sixty percent of I. edulis trees fixed N₂; %N_f was 57% for the N₂-fixing trees. Biomass production and N yield were similar in N₂-fixing and non-N₂-fixing I. edulis. No obvious cause was found for why not all I. edulis trees fixed N₂. All E. poeppigiana and E. fusca trees fixed N₂; %N_f was ca. 59% and 64%, respectively. These data were extrapolated to typical agroforestry systems using published data on N recycling by the studied species. Inga edulis may recycle ca. 100 kg ha⁻¹ a⁻¹ of N fixed from atmosphere to soil if only 60% of trees fix N₂, E. poeppigiana 60–160 kg ha⁻¹ a⁻¹, and E. fusca ca. 80 kg ha⁻¹ a⁻¹.     

CHy-banding patterns and chromatin organization inAegilops andTriticum species (Poaceae)

The distribution of CHy-banded heterochromatin was studied in the chromosomes ofAegilops longissima, Ae. speltoides, Triticum monococcum, andT. turgidum. Interphase nuclei were measured after Feulgen staining at different thresholds of optical density; the curves so obtained indicated the relationship among the species with respect to the different fractions of the genomic DNA. The karyological and cytophotometric analyses indicate differences betweenAe. speltoides andAe. longissima, the latter species being enriched in heterochromatin. Similar results were demonstrated for the genusTriticum, in whichT. turgidum showed more heterochromatin when compared withT. monococcum. The results suggest that the B genome of the cultivated wheats possesses a type of heterochromatin that resembles the type present inAe. longissima.     

Immediate effects of pyridazinone herbicides on photosynthetic electron transport in algal systems

Pyridazinone herbicides, SANDOZ 9785 (4-chloro-5-dimethylamino2-phenyl-3-(2H) pyridazinone), SANDOZ 9789 (4-chloro-5 (methylamino)-2-(α, α, α-trifluoro-m-tolyl-3-(2H) pyridazinone) and SANDOZ 6706 (4-chloro-5-(methylamino)-2-(α, α, α-trifluoro-m-tolyl-3-(2H) pyridazinone) inhibited photosystem II electron transport inChlorella protothecoides, when the herbicides were added to the assay medium. The inhibitory eficiency varied with the algal species and the nature of substitution of pyridazinones. Using 3 algal systemsviz., Chlorella, Scenedesmus andAnacystis, the I₅₀ value of for the inhibition of photosynthesis of 3 substituted pyridazinones (SANDOZ 9785, SANDOZ 6706 and SANDOZ 9789) were determined. SANDOZ 9789 was found to be the weakest inhibitor of photosystem II electron transport (H₂O→ benzoquinone) as compared to SANDOZ 9785 and SANDOZ 6706. In general, the order of inhibition could be given as SANDOZ 6706 >- SANDOZ 9785 > SANDOZ 9789. The I₅₀ value of photosynthetic particles obtained fromChlorella cells was similar to that of whole cells, suggesting that the cell wall ofChlorella did not act as a barrier for the herbicide action. Studies on the light intensity dependence of SANDOZ 9785 inhibition of electron transport (H₂O→ benzoquinone) showed that the light-dependent portion of the curve was more sensitive than the light independent portion of the curve. It is suggested that the site of action was on the reducing side of photosystem II.