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PubMed2006

Tepary bean (Phaseolus acutifolius).

Phaseolus beans are among the major legumes for food consumption, especially in Latin America, Africa, and Asia. Tepary bean (Phaseolus acutifolius L. Gray) is one of the five cultivated species of the genus Phaseolus. This chapter describes an Agrobacterium-mediated transformation protocol for P. acutifolius based on cocultivation of callus, derived from cotyledonary nodes, with Agrobacterium. The selectable marker gene used is neomycin phosphotransferase II (nptII), and the selection agent is geneticin. Selection of transgenic callus material is achieved through four to five passages on geneticin-containing medium, after which shoots are induced on medium without selection agent. The protocol as described here has been applied to transform a cultivated variety of P. acutifolius, TB1, and also with some modifications to a wild genotype, NI576 and another cultivated variety, PI440795.

Agrobacterium tumefaciens↗
PubMed1999

The Tpv2 family of retrotransposons of Phaseolus vulgaris: structure, integration characteristics, and use for genotype classification.

The Tpv2 family of transposable elements of common bean, Phaseolus vulgaris, belongs to the Ty1/copia group of long terminal repeat (LTR) containing retrotransposons. By reverse transcriptase (RT)-PCR and by analysis of genomic clones, we characterized four of the approximately 40 copies present in the Phaseolus genome, and the genomic environment of their integration sites. Tpv2 integrated preferentially into actively transcribed regions. While none of the isolated elements had all the functional domains necessary for transposition, analysis of bean cultivars suggested that some members of the Tpv2 family transposed in recent breeding history. Probes derived from Tpv2, as well as flanking genomic sequences, may be useful for classifying Phaseolus cultivars.

Amino Acid Sequence↗
PubMed1990

Comparison of growth, nitrogen metabolism and organ weights in piglets and rats fed on diets containing Phaseolus vulgaris beans.

The effects of lectins in the diet have been mainly studied in rats. An important question is whether results obtained in rats can be extrapolated to larger animals like the pig. Phaseolus vulgaris beans are rich in toxic lectins. Therefore a study was carried out to compare the effects of diets containing 200 g Phaseolus vulgaris beans (raw or toasted)/kg in rats and piglets. Live-weight gain, nitrogen digestibility and N balance were much lower in piglets than in rats fed on diets containing raw beans. Live-weight gain and N balance were slightly negative in the piglets. When toasted beans were given, live-weight gain and N balance values were reduced in piglets but hardly at all in rats. Giving raw beans caused hypertrophy of the pancreas in the rats but in piglets the weight of the pancreas was reduced. Spleen weight was depressed in the piglets but not in the rats. Weight of liver was not affected in either animal species. When toasted beans were given no effects on the weights of pancreas, spleen or liver were found in piglets or rats. It was concluded that the piglet is much more sensitive to antinutritional factors in the Phaseolus vulgaris bean than the rat.

Animal Feed↗
PubMed1997

Cloning, sequence, and expression of a blood group B active recombinant alpha-D-galactosidase from pinto bean (Phaseolus vulgaris).

A cDNA encoding pinto bean alpha-D-galactosidase [E.C. 3.2.1.22] was obtained by amplification of cDNA using highly conserved sequences found in eucaryotic alpha-D-galactosidases. Subsequently a full length Phaseolus cDNA clone was obtained that is 1537 nt long and contains untranslated 5' and 3' sequences. The nucleotide sequence of the cDNA has a high degree of homology with other eucaryotic alpha-D-galactosidase genes. The recombinant alpha-D-galactosidase (rGal) was expressed in Escherichia coli and purified by ion exchange and affinity chromatography. Purified rGal was homogeneous by SDS-PAGE and had relative masses of 40.1 and 45.4 kDa under nonreducing and reducing conditions, respectively. The N-terminal sequence of the expressed protein contained the sequence GNGLGQTPPMG corresponding to that deduced from the cDNA sequence. The native molecular weight for rGal was determined to be 32.18 kDa by Sephacryl S-200 chromatography. The specific activity of the rGal was 349 mu moles of PNP-alpha-D-galactopyranoside hydrolyzed per mg of pure rGal per min. rGal was highly specific for alpha-D-galactosyl residues and degraded B oligosaccharide. No detectable hemagglutinin or protease activity was present in the preparations. Furthermore, rGal was active against the blood group B antigen on native human erythrocytes in cell suspension assays. The only detectable RBC phenotypic change was loss of the B and P1 epitopes. Recombinant Phaseolus vulgaris alpha-D-galactosidase may have useful biotechnical applications in the potential mass production of enzymatically converted, universally transfusable type O RBCs. alpha-D-galactosidase [E.C. 3.2.1.22] has been purified from a variety of procaryotic and eucaryotic species. Most alpha-D-galactosidases have similar low molecular weight substrate specificities, but activity against high molecular weight substrates is variable. Terminal alpha-D-galactoside residues are present in glycoproteins and glycolipids. Some alpha-D-galactosidases have activity against alpha-D-galactosyl residues on cell membrane glycoconjugates. Glycosidases with this property are useful for carbohydrate structural studies and biotechnical applications. Enzymes free of other glycosidase activities with activity near neutral pH are particularly useful for membrane modification studies on native cells. Complex sugar chains in glycolipids and glycoproteins have often been implicated in the growth and development of eucaryotes. In particular, complex sugar chains play an important role in the recognition of self in the immune system. Some alpha-D-galactosidases can modify certain carbohydrate membrane epitopes, thereby modulating the immune response. For example, the blood group B epitope expressed on erythrocytes contains a terminal alpha-D-galactosyl residue. Individuals lacking this antigen produce naturally occurring complement fixing antibodies to the B epitope. Hydrolysis of this terminal saccharide destroys the antigenic activity of the B determinant producing H antigen (blood type O) on erythrocytes. Only rare individuals produce clinically significant antibodies to the H antigen, and therefore, type O red blood cells are "universally" compatible and in great demand. Dhar purified alpha-D-galactosidase isozymes from Phaseolus vulgaris and characterized their activity. To our knowledge, our laboratory, in a brief report, is the first to describe the cloning of the gene and the use of recombinant enzyme for seroconverting blood type B to O cells. This paper describes the cloning, sequence, expression, purification, and characterization of recombinant alpha-D-galactosidase. Activity of the recombinant enzyme on the native human erythrocyte blood group B epitope is shown.

ABO Blood-Group System↗
PubMed1999

Effect of Atmospheric CO2 Enrichment on Root Growth and Carbohydrate Allocation of Phaseolus spp.

A glasshouse experiment was conducted with plants of Phaseolus grown in liquid culture. Root growth parameters (biomass, diameter, length, growth rate, zone of cell division), root rheological components (wall extensibility, water potential yield threshold, water potential), shoot growth, carbon allocation, and abscisic acid (ABA) concentration were measured in Phaseolus acutifolius A. Gray at ambient (550 µmol mol-1) and elevated (700 µmol mol-1) atmospheric CO2 concentrations. For contrast, measurements of above- and belowground growth were conducted on Phaseolus vulgaris L. in the same treatments. Under nonlimiting conditions of water and nutrients, elevated CO2 increased root and shoot growth of P. acutifolius but not P. vulgaris. While root mass was increased by nearly 60% in P. acutifolius, there was no effect of atmospheric CO2 on any of the rheological components measured. In contrast, starch and ABA accumulated in roots of P. acutifolius. The concentration of starch in roots of P. acutifolius increased by 10-fold, while root concentrations of ABA doubled. From the data it is concluded that CO2 enrichment is favorable for root growth in some species in that more carbon is allocated to belowground growth. In addition, ABA may play a role in growth responses and/or allocation of photosynthates at elevated CO2 in P. acutifolius.

Journal Article↗
PubMed1994

Control of Paraheliotropism in Two Phaseolus Species.

Paraheliotropic (light-avoiding) leaf movements have been associated with high light intensity, high temperature, and drought. We investigated leaf elevation for intact plants, pulvinus bending for excised motor organs, and size change for protoplasts from motor tissue for two Phaseolus species: Phaseolus acutifolius A. Gray, native to hot, arid regions, and Phaseolus vulgaris L., the common bean. Leaf angles above horizontal were measured for central trifoliolate leaflets of intact plants at 24, 27, and 30[deg]C at 500 and 750 [mu]mol photons (400-700 nm) m-2 s-1 over a range of water potentials; equivalent angles were determined for excised motor organs under similar conditions. Diameters were measured for protoplasts from abaxial and adaxial motor tissue over a range of photon flux density values, temperatures, and water potentials. In general, higher photon flux density and temperature resulted in elevation of leaves, bending of excised pulvini, and equivalent changes in protoplast volume (swelling of abaxial protoplasts and shrinking of adaxial protoplasts). In intact plants, lower water potentials yielded greater paraheliotropism; abaxial protoplasts increased in size, whereas adaxial ones did not change. P. acutifolius typically exhibited greater paraheliotropism than did P. vulgaris under the same conditions, a set of physiological responses likely to be highly adaptive in its native arid habitat.

Journal Article↗
PubMed1968

Polar Movement of Indole-3-acetic Acid-C in Roots of Lens and Phaseolus.

A critical review of the few papers on IAA-(14)C movement in roots revealed apparent contradictions, as well as flaws in experimental design that would be apt to cause artifacts. The movement of (14)C from IAA-(14)C was studied in sections of Lens and Phaseolus roots, using a system 20 or more times as sensitive as any previously used. To make sure that our results with roots could be compared validly with published work on petioles and stems, we used the same techniques as we had earlier used for shoot structures. The results with Lens were similar in many ways to those for shoots: net movement into receiver blocks was very strongly polar, followed a linear course for several hours, and showed a velocity of the same order of magnitude as in shoots (and, in fact, very close in absolute value to that found in Coleus stem cylinders). Also, as with shoots, all the radioactivity in receiver blocks ran to the R(F) of IAA. The time-course of loss of counts from donor blocks was similar to that found in shoots. The 2 most striking differences from shoots were 1) the very low percentage of added (14)C that was moved into the receivers (about one-tenth of the values for bean petioles), and 2) the fact that the polar movement was acropetal in roots, rather than basipetal as in shoots. Results with Phaseolus roots were similar to those for Lens, although an additional complication with Phaseolus roots was the indication of a transitory stage of weak basipetal polarity in the first few hours after excising the section. This stage was followed in a few hours by a stronger acropetal polarity.

Journal Article↗
PubMed1977

Purification and initial kinetic characterization of different forms of o-acetylserine sulfhydrylase from seedlings of two species of phaseolus.

Purification of O-acetylserine sulfhydrylase (OASS) from seedlings of two species of Phaseolus reveals the presence in both species of two forms of this enzyme. The isolation and purification procedure gives purification of 7- to 160-fold for individual isoenzymes with specific activities ranging from 33 IU mg(-1) to 775 IU mg(-1) protein.Detailed study of the basic kinetic parameters of the OASS isoenzymes indicates that both forms from Phaseolus vulgaris (which are of about equal specific activity) display substrate inhibition by S(2-) above 1 mm and positive cooperativity at lower concentrations of S(2-). With respect to O-acetylserine (OAS), the second substrate of the reaction, one P. vulgaris isoenzyme shows substrate inhibition by OAS concentrations above 10 mm, while the second is unaffected by OAS concentrations up to 50 mm. The isoenzymes from Phaseolus polyanthus (one of which has a specific activity 24 times higher than the other) are slightly and approximately equally inhibited by both S(2-) and OAS.

Journal Article↗
PubMed1981

Cytokinin-Active Ribonucleosides in Phaseolus RNA: II. DISTRIBUTION IN tRNA SPECIES FROM ETIOLATED P. VULGARIS L. SEEDLINGS.

The distribution of cytokinin-active ribonucleosides in tRNA species from etiolated Phaseolus vulgaris L. seedlings has been examined. Phaseolus tRNA was fractionated by benzoylated diethylaminoethyl-cellulose and RPC-5 chromatography, and the distribution of cytokinin activity was compared with the distribution of tRNA species expected to correspond to codons beginning with U. Phaseolus tRNA(Cys), tRNA(Trp), tRNA(Tyr), a major peak of tRNA(Phe), and a large fraction of tRNA(Leu) were devoid of cytokinin activity in the tobacco bioassay. Cytokinin activity was associated with all fractions containing tRNA(Ser) species and with minor tRNA(Leu) species. In addition, several anomalous peaks of cytokinin activity that could not be directly attributed to U group tRNA species were detected.

Journal Article↗
PubMed1987

Metabolism of C-zeatin in phaseolus embryos : occurrence of o-xylosyldihydrozeatin and its ribonucleoside.

The metabolism of trans-[8-(14)C]zeatin was examined in embryos of Phaseolus acutifolius A. Gray P.I. 321637 and Phaseolus coccineus Lam. cvs Scarlet Runner and Desiree. In both species zeatin was converted to ribosylzeatin, ribosylzeatin 5'-monophosphate, O-glucosyl-9-ribosylzeatin and the recently discovered O-xylosyl derivatives of zeatin and ribosylzeatin (Turner, JE, DWS Mok, MC Mok, G Shaw 1987 Proc Natl Acad Sci USA. In press). Two new metabolites, identified by enzyme degradation and gas chromatography-mass spectrography analyses as O-xylosyldihydrozeatin and its ribonucleoside, were recovered from P. coccineus embryos. From this and previous studies it may be concluded that the potential to form O-xylosyl derivatives of zeatin is present only in embryos of three Phaseolus species (P. vulgaris L., P. coccineus, and P. acutifolius), but not in P. lunatus L., while the reduction of the side chain is most prominent in P. coccineus.

Journal Article↗
PubMed2000

Occupational contact dermatitis to Phaseolus vulgaris in a farmer - a case report.

A case of occupational contact dermatitis in a farmer is described, caused among others by Phaseolus vulgaris. The patient's history of eczematous and vesicular and bullous skin reactions occurring after exposure to Phaseolus was confirmed by skin tests with native leaves of the plant. To the best of our knowledge, this is the first description of occupational contact dermatitis caused by leaves of Phaseolus plant.

Adult↗
PubMed2006

Development of four phylogenetically-arrayed BAC libraries and sequence of the APA locus in Phaseolus vulgaris.

The APA family of seed proteins consists of three subfamilies, in evolutionary order of hypothesized appearance: phytohaemagglutinins (PHA), alpha-amylase inhibitors (alphaAI), and arcelins (ARL). The APA family plays a defensive role against mammalian and insect seed predation in common bean (Phaseolus vulgaris L.). The main locus (APA) for this gene family is situated on linkage group B4. In order to elucidate the pattern of duplication and diversification at this locus, we developed a BAC library in each of four different Phaseolus genotypes that represent presumptive steps in the evolutionary diversification of the APA family. Specifically, BAC libraries were established in one P. lunatus (cv. 'Henderson: PHA+ alphaAI- ARL-) and three P. vulgaris accessions (presumed ancestral wild G21245 from northern Peru: PHA+ alphaAI+ ARL-; Mesoamerican wild G02771: PHA+ alphaAI+ ARL+; and Mesoamerican breeding line BAT93: PHA+ alphaAI+ ARL-). The libraries were constructed after HindIII digestion of high molecular weight DNA, obtained with a novel nuclei isolation procedure. The frequency of empty or cpDNA-sequence-containing clones in all libraries is low (generally <1%). The Henderson, G21245, and G02771 libraries have a 10x genome coverage, whereas the BAT93 library has a 20x coverage to allow further, more detailed genomic analysis of the bean genome. The complete sequence of a 155 kbp-insert clone of the G02771 library revealed six sequences belonging to the APA gene family, including members of the three subfamilies, as hypothesized. The different subfamilies were interspersed with retrotransposon sequences. In addition, other sequences were identified with similarity to chloroplast DNA, a dehydrin gene, and the Arabidopsis flowering D locus. Linkage between the dehydrin gene and the D1711 RFLP marker identifies a potential syntenic region between parts of common bean linkage group B4 and cowpea linkage group 2.

Amino Acid Sequence↗
PubMed2006

Lipid peroxidation in cadmium-treated Phaseolus coccineus plants.

Lipid peroxidation was examined in Phaseolus coccineus plants treated with 25 microM Cadmium (Cd) for 12 days at two different growth stages of the primary leaves. In the leaves of Cd-treated young plants, the content of free fatty acids was higher only after 8 days of plant exposure to the metal, lipoxygenase activity at pH 7.0 was enhanced after 6 and 8 days of Cd action, but at pH 8.0 it was higher than control, especially after 8 days. Non-enzymatic lipid peroxidation was higher after 6 days of young plant exposure to the metal, but after 8 days it slightly decreased. Glutathione-S-transferase activity decreased during 12 days of the metal action but it was always higher than in control. Anthocyanin content in these plants was also elevated in comparison with control. In Cd-treated older plants, the level of free fatty acids was always higher as compared to control. Lipoxygenase activity at pH 7.0 was lower after 4 days of plant exposure to the metal, but then it was close to control. The enzyme activity at pH 8.0 and the level of lipid peroxidation were lower during 12 days of Cd action in comparison with control. Glutathione-S-transferase activity and anthocyanin pigment content were higher in older Cd-treated plants than in control. Relationships between the pool of free fatty acids, a low level of lipid peroxidation, and glutathione-S-transferase activity in the primary leaves of Cd-treated Phaseolus coccineus plants are discussed. In these plants, lipid peroxidation is under the strict control of metabolism and the antioxidative system works efficiently.

Anthocyanins↗
PubMed2006

Agrobacterium tumefaciens mediated transfer of Phaseolus vulgaris alpha-amylase inhibitor-1 gene into mungbean Vigna radiata (L.) Wilczek using bar as selectable marker.

Morphologically normal and fertile transgenic plants of mungbean with two transgenes, bar and alpha-amylase inhibitor, have been developed for the first time. Cotyledonary node explants were transformed by cocultivation with Agrobacterium tumefaciens strain EHA105 harboring a binary vector pKSB that carried bialaphos resistance (bar) gene and Phaseolus vulgaris alpha-amylase inhibitor-1 (alphaAI-1) gene. Green transformed shoots were regenerated and rooted on medium containing phosphinothricin (PPT). Preculture and wounding of the explants, presence of acetosyringone and PPT-based selection of transformants played significant role in enhancing transformation frequency. Presence and expression of the bar gene in primary transformants was evidenced by PCR-Southern analysis and PPT leaf paint assay, respectively. Integration of the Phaseolus vulgaris alpha-amylase inhibitor gene was confirmed by Southern blot analysis. PCR analysis revealed inheritance of both the transgenes in most of the T(1) lines. Tolerance to herbicide was evidenced from seed germination test and chlorophenol red assay in T(1) plants. Transgenic plants could be recovered after 8-10 weeks of cocultivation with Agrobacterium. An overall transformation frequency of 1.51% was achieved.

Agrobacterium tumefaciens↗
PubMed2006

Synergy between the light-induced acute response and the circadian cycle: a new mechanism for the synchronization of the Phaseolus vulgaris clock to light.

PvLHY and Lhcb expression has been studied in primary bean leaves after exposure of etiolated leaves to two or three white light-pulses and under different photoperiods. Under the tested photoperiods, the steady-state mRNA levels exhibit diurnal oscillations with zenith in the morning between ZT21 and 4 for PvLHY and between ZT4 and 6 for Lhcb. Nadir is in the evening between ZT12 and 18 for PvLHY and ZT18 and 24 for Lhcb. Light-pulses to etiolated seedlings induce a differentiated acute response that is reciprocally correlated with the amplitude of the following circadian cycle. In addition, the clock modulates the duration of the acute response (descending part of the curve included), which according to the phase of the rhythm at light application extends from 7 to 18 h. This constitutes the response dynamics of the Phaseolus clock to light. Similarly, the waveform of PvLHY and Lhcb expression during the day of different photoperiods resembles in induction capability (accomplishment of peak after lights-on) and duration (from lights-on phase to trough) the phase-dependent progression of acute response in etiolated seedlings. Consequently, the peak of Lhcb (all tested photoperiods) and PvLHY (in LD 18:6) attained in the photophase corresponds to the acute response peak, while the peak of PvLHY during the scotophase (in LD 12:12 and 6:18) corresponds to the circadian peak. Thus, the effect of the response dynamics in the photoperiod determines the coincidence of the peak with the photo- or scotophase, respectively. This represents a new model mechanism for the adaptation of the Phaseolus clock to light.

Biological Clocks↗
PubMedDate not supplied

Purification and characterization of complex carbohydrate specific isolectins from wild legume seeds: Acacia constricta is (vinorama) highly homologous to Phaseolus vulgaris lectins.

Vinorama isolectins (VL2-VL4) were purified from seeds of Acacia constricta (vinorama) using affinity chromatography on a fetuin-fractogel column followed by cationic-exchange chromatography. Each isolectin fraction presented a characteristic isoelectric point range from 5.5 to 8.4. Under native conditions, VL containing fractions migrated as tetramers of 133 kDa, while in SDS-PAGE, in presence of 2-mercaptoethanol, a single subunit band with M(r) of 34 kDa was observed. VL was found to be a glycoprotein with a 7.5% neutral sugar content. Antibodies to Phaseolus vulgaris lectins PHA and other wild legume lectins as Olneya tesota (palo fierro) PF2 and PF3, and Parkinsonia aculeate (palo verde) PV reacted with VL, but not with anti Glycine max agglutinin SBA or anti Lotus tetragonolobus agglutinin LTA. Furthermore, direct analysis of VL peptides showed sequences homologous to those reported in different lectins of the Phaseolus genus. VL2-VL4 did not have ABO serological or simple sugar specificity, but were inhibited by complex carbohydrates from fetuin and thyroglobulin. Asialofetuin carbohydrates strongly interacted with VL4 and VL3. Vinorama isolectins could be classified as "complex lectins".

Acacia↗
PubMed2005

Biochemical characterization, stability studies and N-terminal sequence of a bi-functional inhibitor from Phaseolus aureus Roxb. (Mung bean).

Herein, we report the purification and biochemical characterization of a novel bi-functional protein proteinase/amylase inhibitor from the dietary leguminous pulse Phaseolus aureus Roxb. (Vigna radiata L.) by means of acetic acid precipitation, salt fractionation, ion-exchange chromatography (DEAE-cellulose) and affinity chromatography on trypsin-sepharose column. P. aureus inhibitor is a bi-functional inhibitor since it exhibits inhibitory activity towards trypsin-like and alpha-chymotrypsin-like serine proteinases as well as against alpha-amylases. It is a helix-rich protein (Mr 13,600) containing approximately eight tyrosines, one tryptophan and two cystines. N-terminal sequence alignment reveals no homology to other proteinase inhibitors reported from Phaseolus sp. thereby confirming that it is a novel inhibitor. Inhibitory activity measurements show that the inhibitor is quite stable even at extremely high temperatures and is only slightly affected by pH changes. Circular dichroism (CD) conformational studies revealed some changes in its near- as well as far-ultraviolet spectrum at extremes of pH and temperature. Treatments with trypsin for varying time periods did not alter its proteolytic inhibitory activity but caused some reduction in its amylase inhibitory activity.

Amylases↗
PubMed2004

Performance of phaseolus bean rhizobia in soils from the major production sites in the Nile Delta.

The symbiotic and competitive performances of two highly effective rhizobia nodulating French bean P. vulgaris were studied in silty loam and clayey soils. The experiments were carried out to address the performance of two rhizobia strains (CE3 and Ph. 163] and the mixture thereof with the two major cultivated bean cultivars in two soil types from major growing French bean areas in Egypt. Clay and silty loam soils from Menoufia and Ismailia respectively were planted with Bronco and Giza 6 phaseolus bean cultivars. The data obtained from this study indicated that rhizobial inoculation of Giza 6 cultivar in clayey soil showed a positive response to inoculation in terms of nodule numbers and dry weight. This response was also positive in dry matter and biomass accumulation by the plants. The inoculant of strain CE3 enhanced plant growth and N-uptake relative to Ph. 163. However, the mixed inoculant strains were not always as good as single strain inoculants. The competition for nodulation was assessed using two techniques namely fluorescent antibody testing (FA) and REP-PCR fingerprinting. The nodule occupancy by inoculant strain Ph. 163 in both soils occupied 30-40% and 38-50 of nodules of cultivar Bronco. The mixed inocula resulted in higher proportions of nodules containing CE3 in silty loam soil and Ph. 163 in clayey soil. The native rhizobia occupied at least 50% of the nodules on the Bronco cultivar. For cultivar Giza 6, the native rhizobia were more competitive with the inoculant strains. Therefore, we suggest using the studied strains as commercial inocula for phaseolus bean.

DNA Fingerprinting↗