Comparative study of seed albumins in the Old-WorldLupinus species (Fabaceae) by reversed-phase HPLC

Seed albumins and 2S proteins isolated from the albumin fraction of 36 accessions representing 10 Old-WorldLupinus species (5 smooth- and 5 rough-seeded) were studied using reversed-phase high-performance liquid chromatography. In addition, the globulin fraction was analyzed to determine its 2S protein content. The performed separations showed the suitability of RP-HPLC technique in the analysis of variation of the seed albumin composition in lupins. In the group of rough-seeded lupins, 3 types of RP-HPLC elution profiles of albumins were distinguished: (1)L. atlanticus, (2)L. cosentinii andL. digitatus, (3)L. palaestinus andL. pilosus. All the species of this group were found to have proteins not observed in smooth-seeded species. Smooth-seeded species exhibited more abundant protein spectra, each species distinguishing by its specific RP-HPLC elution profile. It was found that 2S proteins classified as 2S albumins were responsible for the observed variation. Depending onLupinus species, the 2S albumin class consists of two to six proteins.     

Molecular phylogeny and phylogeography ofLupinus (Leguminosae) inferred from nucleotide sequences of therbcL gene and ITS 1 + 2 regions of rDNA

Total DNA was extracted from 55 species of theLeguminosae (including 29 species ofLupinus). The chloroplast generbcL and the ITS 1 + 2 regions of nuclear RNA genes were amplified by polymerase chain reaction (PCR) and sequenced directly. The sequences obtained were evaluated with character state (Maximum Parsimony) and distance methods (Neighbour Joining). Phylogenetic trees obtained with both data sets and methods are mostly congruent.Genisteae andCrotalarieae are sister groups and share ancestry with theThermopsideae/Podalyrieae. The genusLupinus, which forms a monophyletic clade within theGenisteae, shows a distinct Old-New World disjunction and appears to be divided into several more or less distinct groups: (1) The species from the eastern part of South America. (2) The homogeneous rough-seeded group (Scabrispermae) of the Old World species which is well distinguished from the smooth-seeded group (Malacospermae). (3) Within the rather heterogeneous smooth-seeded lupins a smaller subgroup withL. angustifolius, L. hispanicus andL. luteus is recognized. (4) Also separated are North American lupins and South American species with a western distribution. Genetic distances imply that the genusLupinus evolved during the last 12–14 million years, ruling out the hypothesis that the present Old-New World disjunction can be interpreted as a result of the continental drift. The genetic data suggest an origin in the Old World and an independant colonisation of the Eastern parts of South America as opposed to North America and the Western parts of South America.     

A serological contribution to the systematics of the genusLupinus (Fabaceae)

The immunological comparison of the seed reserve proteins suggests thatLupinus is a natural genus, the American and the Euro-African species belonging to the same stock. Among the Lupines of the Old World, the smooth-seeded and the rough-seeded species from two natural segregates. The genusLupinus is serologically related to the rest of theGenisteae and to the AsiaticSophoreae rather than to AmericanSophoreae andThermopsideae. The data suggest thatLupinus may have originated with the remainder of theGenisteae from primitiveSophoreae of temperate-subtropical Asia. America and the Mediterranean-African region are regarded as secondary centres of speciation.     

Electrophoretic seed albumin patterns inVicia species of sectt.Hypechusa andPeregrinae (Fabaceae)

This work is a continuation of electrophoretic investigations aimed at revealing a wild relative ofVicia faba. Electrophoretic analysis (PAGE) of seed albumins covered 52 accessions representing eightVicia species of sect.Hypechusa and two species of sect.Peregrinae. Most of the examined species showed an intraspecific variation due to differences between accessions and/or individual variation within accessions. In spite of the intraspecific variation, marked interspecific differences were recorded. However, none of the investigated species displayed electrophoretic seed albumin patterns similar to those reported earlier forV. faba. Contribution of the obtained results to characterization of the examined taxa is discussed.     

Electrophoretic seed albumin patterns and species relationships inVicia sect.Faba (Fabaceae)

Electrophoretic analysis of seed albumins (PAGE) covered 173 accessions representing nine species ofVicia sect.Faba. The number of albumin bands recorded in particular species varied from three inV. eristaloides to 23 inV. faba; in total, 38 bands were distinguished in the investigated material. The examined species, exceptV. eristalioides, showed intraspecific variation with respect to the number and relative staining intensity of albumin bands; individual variation was especially marked inV. faba and inV. narbonensis. Hierarchical clustering of the investigated taxa was based onBhattacharyya distances calculated from the electrophoretic data. The taxa grouped in three main clusters.Vicia faba and the rather remotely relatedV. kalakhensis formed one cluster. The second cluster was composed ofV. narbonensis distantly related toV. hyaeniscyamus. The third cluster comprised three subgroups: 1.V. johannis, V. galilaea andV. serratifolia, 2.V. eristalioides, and 3.V. bithynica. The obtained results are discussed with reference to taxonomic relationships inVicia sect.Faba.     

Comparative HPLC analysis of seed albumins fromVicia faba andV. kalakhensis (Fabaceae)

Previously reported electrophoretic seed albumin data have shown an unexpected association ofVicia faba withV. kalakhensis. In the present work, seed albumins ofV. faba (subsp.paucijuga and subsp.faba) were compared with those ofV. kalakhensis using ionexchange (IE) and reversed-phase (RP) high-performance liquid chromatography (HPLC). Two subspecies ofV. faba displayed similar seed albumin profiles. On the other hand, seed albumin profiles ofV. faba andV. kalakhensis showed no major protein peak in common either in IE-HPLC or RP-HPLC chromatograms. The reported differences in seed albumin composition ofV. faba andV. kalakhensis are consistent with other taxonomical data showingV. faba to be genetically distant from the wild relatives.     

A comparative electrophoretic study of the seed albumins fromSophora microphylla andPisum sativum (Leguminosae)

The albumin proteins from seed ofSophora microphylla Ait. and from cotyledons ofPisum sativum L. (cv. Greenfeast) have been analysed electrophoretically using a range of gels of varied pore size. Plots of mobility [as 100 log₁₀ (R _f × 100)] vs.acrylamide content of gel indicate that very few of the albumins fromS. microphylla are homologous with albumins fromP. sativum. Despite the diverse compositions of the two fractions, their amino acid analyses were surprisingly similar.     

Genomic relationships among 11 species in the genus Cajanus as revealed by seed protein (albumin and globulin) polymorphisms

SDS-PAGE analysis of seed albumins and globulins from two pigeonpea, Cajanus cajan, cultivars (DSLR-17 and BDN-2) and ten wild species, including C. cajanifolius, C. lineatus, C. sericeus, C. acutifolius, C. lanceolatus, C. reticulates, C. albicans, C. scarabaeoides, C. volubilis and C. platycarpus, resulted in 34 albumin and 27 globulin polypeptides. Proximity matrix analysis based on electrophoretic banding patterns of albumins and globulins jointly revealed C. cajanifolius to be closest to C. cajan having similarity coefficients of 0.595 and 0.676, respectively. Cluster analysis also exhibited the grouping of C. cajanifolius with C. cajan in one cluster. C. platycarpus has been out grouped. The present analysis more or less agreed with the sectional classification of the genus Cajanus and it has been hypothesized that cultivated pigeonpea has evolved through multi-genomic interaction involving C. cajanifolius and that it has experienced minor genomic reorganization during its divergence.     

Hair morphology and the relationships of species inSolanum sect.Basarthrum

The usefulness of features of leaf hairs in distinguishing subgenera and sections is well documented in bothRhododendron andSolanum. In this analysis of the taxa ofSolanum sect.Basarthrum (23 species), and of a sample of closely related taxa from sect.Petota (22 species), it is shown that such features serve to delineate subsectional groups such as series and some species as well. SectionBasarthrum has an unexpected diversity of hair types. Although this group has been characterized by 2-celled bayonet hairs, more than one half of the taxa in the section bear multicellular finger hairs, and 3 species also possess branched hairs. Thus, major rearrangements of the species previously assigned to sect.Basarthrum are indicated or supported by pubescence features. The taxa studied from seriesEtuberosa andJuglandifolia (both of sect.Petota) show hair types that a) are relatively primitive for the section, and b) show linkage between sects.Petota andBasarthrum.     

Functional characterisation of two cytochrome<Emphasis Type="Italic"> b</Emphasis><Subscript>5</Subscript>-fusion desaturases from<Emphasis Type="Italic"> Anemone leveillei</Emphasis>: the unexpected identification of a fatty acid Δ<Superscript>6</Superscript>-desaturase

The Ranunculaceae are known to accumulate a wide range of unusual fatty acids in their seed lipids, and this variability has been advocated as a taxonomic marker. The Anemone species, Anemone leveillei L. and Anemone rivularis Buch.-Ham., have previously been reported to accumulate ⁵-desaturated fatty acids in their seed tissue [K. Aitzetmüller (1995) Plant Syst Evol 9:229–240]. Two cDNAs, AL1 and AL2, with similarity to plant cytochrome b₅-fusion "front-end" desaturases were isolated from developing seeds of A. leveillei and their function identified by expression in Saccharomyces cerevisiae. AL2 was characterised as a sphingolipid long-chain-base ⁸-desaturase, while AL1 acted as a fatty acid desaturase. However, AL1 did not produce ⁵-desaturated fatty acids as expected; instead, when expressed in transgenic S. cerevisiae or Arabidopsis thaliana this enzyme was functionally characterised as a ⁶-desaturase. Northern analysis confirmed the expression of this gene in seed tissue and leaf tissue of A. leveillei, though ⁶-desaturated fatty acids were found to accumulate only in the leaf tissue. The unexpected characterisation of a ⁶-desaturase in A. leveillei has implications for the use of fatty acids in chemotaxonomic studies. This is also the first report of a higher-plant ⁶-desaturase from a family other than the Boraginaceae.Abbreviations ALA -linolenic acid - DMOX 4,4-dimethyloxazoline - EDA eicosadienoic acid - FAME fatty acid methyl ester - GLA -linolenic acid - LA linoleic acid - LCB long chain base - ORF open reading frame - OTA octadecatetraenoic acid     

The β-tubulin gene family evolution in theDrosophila montium subgroup of themelanogaster species group

The 1-, 2-, and 3-tubulin genes have been mapped by in situ hybridization on the polytene chromosomes of 11 selected species (15 strains) belonging to theDrosophila montium subgroup. Although the hybridization pattern among the strains of the same species does not differ, this pattern is significantly different among the species. The -tubulin genes in themontium subgroup seem to be organized in a cluster, or in a semi-cluster, or are completely dispersed. The clustered arrangement is found in the North-Oriental sibling speciesD. auraria, D. triauraria, andD. quadraria. The semi-clustered arrangement, wherein the 1 and 2 genes are located at the same locus while 3 is at a different one, appears in the South-Oriental speciesD. bicomuta, D. serrata, andD. birchii, as well as in the Afrotropical speciesD. diplacantha andD. seguyi. The complete separation of the genes is observed in the Indian speciesD. kikkawai andD. jambulina and in the Afrotropical speciesD. vulcana. Based on the above results, a possible mode of evolution of the -tubulin genes in the montium subgroup is attempted. In addition, phylogenetic relationships among themontium species are discussed. Correspondence to: Z.G. Scouras     

Hybridization between South American Jarilla and North American diploid creosotebush (Larrea,Zygophyllaceae)

Different strains of the two diploidLarrea speciesL. divaricata (S. America) andL. tridentata (N. America), are compared morphologically, cytologically, and in their seed proteins. There are minor differences, also in chiasma frequencies. Experimental F₁ hybrids have near-normal meiosis but are semisterile. The parental species, therefore, biologically may be considered allopatric semispecies.     

Alkaloids of theSedum acre-group (Crassulaceae)

The 16 species of theSedum acre-group were investigated for the presence of alkaloids. They areS. acre ofS. ser.Acria, S. alpestre, S. annuum, S. apoleipon, S. borissovae, S. euxinum, S. grisebachii, S. laconicum, S. multiceps, S. sexangulare, S. tuberiferum, S. tuberosum, S. ursi, andS. urvillei ofS. ser.Alpestria, S. samium ofS. ser.Samia, andS. litoreum ofS. ser.Litorea. S. acre differs significantly from the other species. It contains sedamine, hydroxy sedamine, and a number of 2,6-disubstituted piperidine alkaloids. The leafy parts of the species ofS. ser.Alpestria, S. ser.Samia, andS. ser.Litorea contain 4 piperidine alkaloids which also occur inS. acre, and in addition 4 pyrrolidine alkaloids not present inS. acre. The composition of the alkaloid fraction agrees with the infrageneric classification (series) based on the hybridization patterns of the species (comparia).     

Biochemical and immunocytochemical identification of conglutin γ, a lupin seed lectin,in the roots of young germinating seedlings

Summary Polyclonal antibodies directed against polypeptide epitopes of conglutin , a glycosylated lectin in lupin seed, have been used to identify and quantify this protein in root extracts of germinating lupins. The highest conglutin content was found in protein extracts of root elongation zones after 5 to 7 days germination. Root conglutin showed the same subunit composition, glycosylation pattern, isoelectric point, and lectin activity as the cotyledonary one. Immunolocalization experiments on root thin sections demonstrated that conglutin is chiefly present in the intercellular spaces of the cortical parenchyma, where it forms large aggregates. Labelling of the Golgi complexes and the area between the plasmalemma and cell wall revealed the conglutin pathway from post-synthetic processing to excretion via the secretory system.Abbreviations EDTA ethylene diamino tetraacetic acid - IEF isoelectric focusing - LRW London resin white - NC nitrocellulose - PAGE polyacrylamide gel electrophoresis - pI isoelectric point - P_i inorganic phosphate - SDS sodium dodecylsulphate - TEM transmission electron microscopy     

β-Thalassaemia/haemoglobin E tissue ferritins

Summary Ferritins from liver and spleen of both-thalassaemia/haemoglobin E (HbE) and non-thalassaemic patients were purified by heating a methanol-treated homogenate, followed by molecular exclusion chromatography. The concentrations of ferritins in the-thalassaemia/HbE liver and spleen were calculated as 3.8 and 2.0 mg/g wet tissue. The-thalassaemia/HbE ferritin iron/protein ratios were higher than those of normal ferritins. On PAGE, all ferritins gave a single major monomeric band with only very small differences in their mobility. Ferritins from thalassaemic patients also possessed bands corresponding to oligomers. On SDS/PAGE, all ferritins were resolved into two major subunits: H and L with L subunit predominating. While the isoferritin profiles of ferritins from-thalassaemia/HbE liver and spleen were similar to each other and to those of normal liver and spleen, some extra bands were present in the acidic region. The microstructure of these pathological ferritins appears to result, to a large degree, from the particular nature and amount of iron loading present.Abbreviations PAGE polyacrylamide gel electrophoresis - SDS sodium dodecyl sulphate     

Production of 22:2<Superscript>Δ5,Δ13</Superscript> and 20:1<Superscript>Δ5</Superscript> in <Emphasis Type="Italic">Brassica carinata</Emphasis> and soybean breeding lines via introduction of <Emphasis Type="Italic">Limnanthes</Emphasis> genes

Seed oils of meadowfoam (Limnanthes douglasii, L. alba) contain very long-chain fatty acids of strategic importance for a number of industrial applications. These include the monoene 20 1⁵ and the diene 22:2^5,13. Engineering of meadowfoam-type oils in other oilseed crops is desirable for the production of these fatty acids as industrial feedstocks. Accordingly, we have targeted Brassica carinata and soybean (Glycine max) to trangenically engineer the biosynthesis of these unusual fatty acids. An L. douglasii seed-specific cDNA (designated Lim Des5) encoding a homolog of acyl-coenzyme A desaturases found in animals, fungi and cyanobacteria was expressed in B. carinata, which resulted in the accumulation of up to 10% 22:2^5,13 in the seed oil. In soybean, co-expression of Lim Des5 with a cDNA (Lim FAE1) encoding an FAEl (elongase complex condensing enzyme) homolog from L. douglasii resulted in the accumulation of 20:1⁵ to approximately 10% of the total fatty acids of seeds. The content of C₂₀ and C₂₂ fatty acids was also increased from <0.5% in non-transformed soybean seeds to >25% in seeds co-expressing the Lim. douglasii Des5 and FAE1 cDNAs. In contrast, expression of the Lim Des5 in Arabidopsis did not produce the expected 20:2^5,11 in the seed oil. Cumulatively, these results demonstrate the utility of soybean and B. carinata for the production of vegetable oils containing novel C₂₀ and C₂₂ fatty acids, and confirm that the preferred substrates of the Lim Des5 are 20:0 and 22:1³, respectively.     

Taxonomic evaluations of the genusPinus (Pinaceae) based on electrophoretic data of salt soluble and insoluble seed storage proteins

Electrophoretic studies on seed storage proteins of various gymnosperm species showed that both salt soluble and insoluble protein fractions give taxonomic information. Among species of the genusPinus evident differences are found between the subgeneraHaploxylon (Strobus) andDiploxylon (Pinus). In sect.Strobus the two subsectt.Strobi andCembrae are readily distinguished fromP. bungeana (sect.Parrya) which appears to fall in an intermediate position between haploxyl and diploxyl pines. Among the species of subg.Diploxylon, mountain pines show very similar protein patterns in agreement with their recent speciation (Quaternary).     

In vitro physiological approach to classification ofFrankia isolates of ‘the Alnus group’, based on urease,protease and β-glucosidase activities

Summary Most of theFrankia strains isolated fromAlnus andMyrica species are morphologically almost indistinguishable, when grown under standard culture conditions. They form similar vegetative hyphae while sporangia are produced in variable amounts from strain to strain.Physiological reactions were assessed in order to compare 20 strains isolated from various species ofAlnus and one species ofMyrica in Europe and North America. Among invariant negative or positive characteristics, differences in urease, protease and -glucosidase activities appeared to be of significant value.