SEARCH · PubMed Health

Results for “ALFALFA”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 793 records · Page 44Linked to original sources

PubMed2005

Coat protein activation of alfalfa mosaic virus replication is concentration dependent.

Alfalfa mosaic virus (AMV) and ilarvirus RNAs are infectious only in the presence of the viral coat protein; therefore, an understanding of coat protein's function is important for defining viral replication mechanisms. Based on in vitro replication experiments, the conformational switch model states that AMV coat protein blocks minus-strand RNA synthesis (R. C. Olsthoorn, S. Mertens, F. T. Brederode, and J. F. Bol, EMBO J. 18:4856-4864, 1999), while another report states that coat protein present in an inoculum is required to permit minus-strand synthesis (L. Neeleman and J. F. Bol, Virology 254:324-333, 1999). Here, we report on experiments that address these contrasting results with a goal of defining coat protein's function in the earliest stages of AMV replication. To detect coat-protein-activated AMV RNA replication, we designed and characterized a subgenomic luciferase reporter construct. We demonstrate that activation of viral RNA replication by coat protein is concentration dependent; that is, replication was strongly stimulated at low coat protein concentrations but decreased progressively at higher concentrations. Genomic RNA3 mutations preventing coat protein mRNA translation or disrupting coat protein's RNA binding domain diminished replication. The data indicate that RNA binding and an ongoing supply of coat protein are required to initiate replication on progeny genomic RNA transcripts. The data do not support the conformational switch model's claim that coat protein inhibits the initial stages of viral RNA replication. Replication activation may correlate with low local coat protein concentrations and low coat protein occupancy on the multiple binding sites present in the 3' untranslated regions of the viral RNAs.

Alfalfa mosaic virus↗

PubMed1997

Identification of a competitive translation determinant in the 3' untranslated region of alfalfa mosaic virus coat protein mRNA.

We report that the competitive translational activity of alfalfa mosaic virus coat protein mRNA (CP RNA), a nonadenylated mRNA, is determined in part by the 3' untranslated region (UTR). Competitive translation was characterized both in vitro, with cotranslation assays, and in vivo, with microinjected Xenopus laevis oocytes. In wheat germ extracts, coat protein synthesis was constant when a fixed amount of full-length CP RNA was cotranslated with increasing concentrations of competitor globin mRNA. However, translation of CP RNA lacking the 3' UTR decreased significantly under competitive conditions. RNA stabilities were equivalent. In X. laevis oocytes, which are translationally saturated and are an inherently competitive translational environment, full-length CP RNA assembled into large polysomes and coat protein synthesis was readily detectable. Alternatively, CP RNA lacking the 3' UTR sedimented as small polysomes, and little coat protein was detected. Again, RNA stabilities were equivalent. Site-directed mutagenesis was used to localize RNA sequences or structures required for competitive translation. Since the CP RNA 3' UTR has an unusually large number of AUG nucleotide triplets, two AUG-containing sites were altered in full-length RNA prior to oocyte injections. Nucleotide substitutions at the sequence GAUG, 20 nucleotides downstream of the coat protein termination codon, specifically reduced full-length CP RNA translation, while similar substitutions at the next AUG triplet had little effect on translation. The competitive influence of the 3' UTR could be explained by RNA-protein interactions that affect translation initiation or by ribosome reinitiation at downstream AUG codons, which would increase the number of ribosomes committed to coat protein synthesis.

Alfalfa mosaic virus↗

PubMed2004

Spatial determinants of the alfalfa mosaic virus coat protein binding site.

The biological functions of RNA-protein complexes are, for the most part, poorly defined. Here, we describe experiments that are aimed at understanding the functional significance of alfalfa mosaic virus RNA-coat protein binding, an interaction that parallels the initiation of viral RNA replication. Peptides representing the RNA-binding domain of the viral coat protein are biologically active in initiating replication and bind to a 39-nt 3'-terminal RNA with a stoichiometry of two peptides: 1 RNA. To begin to understand how RNA-peptide interactions induce RNA conformational changes and initiate replication, the AMV RNA fragment was experimentally manipulated by increasing the interhelical spacing, by interrupting the apparent nucleotide symmetry, and by extending the binding site. In general, both asymmetric and symmetric insertions between two proposed hairpins diminished binding, whereas 5' and 3' extensions had minimal effects. Exchanging the positions of the binding site hairpins resulted in only a moderate decrease in peptide binding affinity without changing the hydroxyl radical footprint protection pattern. To assess biological relevance in viral RNA replication, the nucleotide changes were transferred into infectious genomic RNA clones. RNA mutations that disrupted coat protein binding also prevented viral RNA replication without diminishing coat protein mRNA (RNA 4) translation. These results, coupled with the highly conserved nature of the AUGC865-868 sequence, suggest that the distance separating the two proposed hairpins is a critical binding determinant. The data may indicate that the 5' and 3' hairpins interact with one of the bound peptides to nucleate the observed RNA conformational changes.

Alfalfa mosaic virus↗

PubMed2005

Detection of Alfalfa mosaic virus (AMV) in pea field in Iran.

During the spring and summer, in 2003-2004, pea viruses were identified in twenty pea fields of Tehran. Some leaf samples were collected randomly from pea fields of Tehran. Samples were tested by Double Antibody Sandwich Enzyme Linked Immunosorbent Assay (DAS-ELISA) technique using polyclonal antiserum of Alfalfa mosaic virus (AMV), AS-0001, DSMZ, Braunschweig, Germany). The samples were extracted in 0.1 M Phosphate buffer pH 7 to 7.5 and inoculated on Chenopodium amaranticolor, Chenopodium quina, Phaseolus valgaris, Vicia faba, Vignia unguiculata. Pea cultivars were infected by AMV, causing mild mosaic, translucent veins and a diffuse green-yellow of tender parts and spots may also was involved necrosis of tissue. Infected plants grow slowly and malformed pods produce fewer ovules. In Chenopodium amranticolor, C. quina chlorotic and necrotic flecks, and Vicia faba systemic mosaic had produced. Phaselous vulgaris and Viginia unguiculata are good assay hosts for strains that produce local lesions after 3-5 days in these plants. Back inoculated on Pisum sativum and Vicia faba and tested with DAS-ELISA that had been confirmed the results. This is the first report of AMV on pea from Iran.

Alfalfa mosaic virus↗

PubMed1993

Construction and characterization of monoclonal antibodies to alfalfa mosaic virus.

Seven monoclonal antibodies (MoAbs) to alfalfa mosaic virus (AMV) isolate T6 were prepared and characterized. One of them (amv 4) reacted with the cryptotope, another (amv 5) with the neotope and the remaining five with various metatopes. Five MoAbs derived from mouse ascitic fluid reacted in immunodiffusion test with native virus and two MoAbs with AMV coat protein. In competitive ELISA, homologous MoAbs caused 80-100% inhibition of the reaction, but some reactions were also affected by heterologous MoAbs. The MoAbs raised against AMV isolate T6 could not be used for differentiation of the four AMV pathotypes examined.

Alfalfa mosaic virus↗

PubMed1995

Characterization of the alfalfa mosaic virus strain T6.

A strain T6 of alfalfa mosaic virus (AlMV) was characterized. It was isolated from field grown lucerne. Purified virus preparations contained four types of particles, B, M, Tb and Ta, containing separately encapsidated ssRNAs 1 to 4. The strain T6 was able to infect 40 different plant species of 9 families, and to develop a systemic infection in most of them. The symptomatology on bean and the RNA mobility of the AlMV strains T6 and 425 were compared. The classical cross-protection experiments on bean have shown that plants inoculated with strain 425 did not develop symptoms of the challenge strain T6.

Alfalfa mosaic virus↗

PubMed1995

Effect of hydrocarbons and crude oil contamination on the sensitivity of French bean to alfalfa mosaic virus.

Determination of local necrotic lesions on primary leaves infected by alfalfa mosaic virus (AMV) revealed that hydrocarbons (HC) contamination of the substrate used for cultivation of French bean (Phaseolus vulgaris L., cv. Black Turtle Soup) caused a reduction of bean leaf area and an increase of plant sensitivity to AMV infection. On the other hand, superficial contamination of the leaves by crude oil caused an inhibition of lesion formation. Changes of SDS-polyacrylamide gel electrophoresis (SDS-PAGE) patterns of extractable bean leaf proteins related to the cultivation substrate contamination by HC were also detected.

Alfalfa mosaic virus↗

PubMed1996

Alfalfa mosaic virus strains T6 and 425 differ in their capsid protein.

Capsid proteins (CPs) of two strains of alfalfa mosaic virus (AIMV)-T6 and 425-were compared using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) peptide mapping and two-dimensional PAGE (2D-PAGE). The CPs had identical molecular mass but differed in their peptide pattern and charge. The AIMV strain T6 was isolated from lucerne in Czech Republic (Gallo, 1977). Previously, its morhology, symptomatology and RNA electrophoretic mobility were characterized and compared with those of the strain 425 (Hagedorn and Handson, 1963; Kúdela and Gallo, 1995). In this study we show that the CP composition of these two AIMV strains differs, too. AIMV, the unique member of the genus Alfamovirus, family Bromoviridae, is of economic significance in many countries in the world, particularly for its seed transmissibility and relative broad host range. Plenty of strains with different biological properties have been isolated (Van Regenmortel and Pinck, 1981). Some of them have been shown to differ in their RNA non-coding regions or in CP composition (Kraal, 1975; Kraal et al., 1976; Collot et al., Dore et al., 1989; Neeleman et al., 1991).

Alfalfa mosaic virus↗

PubMed1999

Production of a diagnostic monoclonal antibody in perennial alfalfa plants.

The increasing use of monoclonal antibodies (mAbs) in diagnostic reagents necessitates efficient and cost-effective mAb production methods. In blood banks, one of the most routinely used reagents is the anti-human IgG reagent used for the detection of non-agglutinating antibodies. Here we report the production of a functional, purified anti-human IgG, through the expression of its encoding genes in perennial transgenic alfalfa. Transgenic plants expressing the light- and heavy-chain encoding mRNAs were obtained, and plants from crosses were found to express fully assembled C5-1. The purification procedure yielded mainly the H2L2 form with specificity and affinity identical to those of hybridoma-derived C5-1. The ability to accumulate the antibody was maintained both in parental F1 lines during repeated harvesting and in clonal material; the antibody was stable in the drying hay as in extracts made in pure water. Also, plant and hybridoma-derived C5-1 had similar in vivo half-lives in mice. These results indicate that plant C5-1 could be used in a diagnostic reagent as effectively as hybridoma-derived C5-1, and demonstrates the usefulness of perennial systems for the cost-effective, stable, and reliable production of large amounts of mAbs.

Animals↗

PubMed2004

Characterization of triacylglycerols from overwintering prepupae of the alfalfa pollinator Megachile rotundata (Hymenoptera: Megachilidae).

Alfalfa leafcutting bees, Megachile rotundata (F.), overwinter as prepupae. The internal lipids were extracted from prepupae that had been wintered at 4 degrees C for 7 months. Megachile rotundata prepupae possessed copious quantities of internal lipids (20% of the fresh weight) that were extracted with CHCl3/methanol (2:1). Transmission electron microscopy revealed that lipids were stored within very large intracellular vacuoles. Separation by silica chromatography revealed that 88% of the internal lipids were triacylglycerols. Ester derivatives of fatty acids from triacylglycerol components were analyzed by gas chromatography-mass spectrometry and 15 fatty acid constituents were identified. The majority (76%) of the triacylglycerol fatty acids were unsaturated fatty acids. The major triacylglycerol fatty acid constituent (30%) was the C16 monounsaturated fatty acid, palmitoleic acid (16:1, hexadec-9-enoic acid), with substantial amounts of linolenic acid (18:3, octadec-9,12,15-trienoic acid, 15%), palmitic acid (16:0, hexadecanoic acid, 14%) and oleic acid (18:1, octadec-9-enoic acid, 13%). Palmitoleic acid as the major fatty acid of an insect is an unusual occurrence as well as the presence of the 16-carbon polyunsaturated fatty acids, 16:2 and 16:3. The major intact triacylglycerol components were separated and identified by high performance liquid chromatography-mass spectrometry. A complex mixture of approximately 40 triacylglycerol components were identified and major components included palmitoyl palmitoleoyl oleoyl glycerol, palmitoyl palmitoleoyl palmitoleoyl glycerol, myristoyl palmitoleoyl palmitoleoyl glycerol, myristoleoyl palmitoyl palmitoleoyl glycerol, and palmitoyl palmitoleoyl linolenoyl glycerol. The function of these internal lipids and their relevance to winter survival and post-wintering development of M. rotundata is discussed.

Animals↗

PubMed1987

Application of polyelectrolytes to fractionation of alfalfa juice protein (short communication).

The quantities of protein which can be extracted from green plants depend on a number of factors such as the botanical composition of the plant, its growth stage, topdressing and system of extraction. Leaf protein concentrates have been incorporated into food for human consumption, but their green color has limited its application. Thermal precipitation of chloroplastic material is not specific, causes denaturation and requires high energy input. A selective recovery of the green fraction is possible by centrifugation at high centrifugal forces or by the combination of this method with an addition of calcium salt to a green juice. Application of high molecular polyelectrolytes in green juice precipitation allows the separation of pigmented material at room temperature. The objective of this work was to determine the effect of cationic and anionic flocculants on fractionation of alfalfa green juice into two fractions: chloroplastic (green LPC) and cytoplasmatic (white LPC).

Chemical Fractionation↗

PubMed1987

Expression of alfalfa mosaic virus coat protein gene confers cross-protection in transgenic tobacco and tomato plants.

A chimeric gene encoding the alfalfa mosaic virus (AlMV) coat protein was constructed and introduced into tobacco and tomato plants using Ti plasmid-derived plant transformation vectors. The progeny of the self-fertilized transgenic plants were significantly delayed in symptom development and in some cases completely escaped infection after inoculated with AlMV. The inoculated leaves of the transgenic plants had significantly reduced numbers of lesions and accumulated substantially lower amounts of coat protein due to virus replication than the control plants. These results show that high level expression of the chimeric viral coat protein gene confers protection against AlMV, which differs from other plant viruses in morphology, genome structure, gene expression strategy and early steps in viral replication. Based on our results with AlMV and those reported earlier for tobacco mosaic virus, it appears that genetically engineered cross-protection may be a general method for preventing viral disease in plants.

Journal Article↗

PubMed1987

Expression of alfalfa mosaic virus RNA 4 in transgenic plants confers virus resistance.

Agrobacterium-mediated transfer from a binary vector was used to produce transgenic Nicotiana tabacum plants that expressed coat protein of the plant virus, alfalfa mosaic virus (AMV). Expression levels of the chimeric gene, which was under the control of the cauliflower mosaic virus 19S promoter, were determined in primary transformed plants, in the progeny from self-fertilization and in the progeny from crosses to normal tobacco. RNA transcripts that were of the expected size as well as a protein of the M(r) and antigenicity of AMV coat protein accumulated in the transgenic plants. Plants that expressed the highest levels of coat protein developed fewer primary infections following inoculation with two strains of AMV and developed systemic infection slower than did plants that did not express coat protein. Resistance was specifically against virions of the AMV strains. AMV RNA and the unrelated virus, tobacco mosaic virus, were as infectious on progeny that expressed coat protein as they were on progeny that did not. The relationship between the virus resistance expressed by these transgenic plants and that observed in virus cross-protection is discussed.

Journal Article↗

PubMed1996

Use of RAPD analysis for in situ identification of Ascosphaera aggregata and Ascosphaera larvis in larval cadavers of the alfalfa leafcutting bee, Megachile rotundata.

Chalkbrood of the alfalfa leafcutting bee, Megachile rotundata, is caused by the fungus Ascosphaera aggregata. We used random amplified polymorphic DNA (RAPD) analysis for the in situ identification of A. aggregata and a related species, Ascosphaera larvis, in larval cadavers of M. rotundata. A simple DNA extraction method was developed to preferentially isolate DNA from fungal spores on the cadaver surface, or from ascocysts beneath the cuticle. Similar banding patterns were obtained in A. aggregata-infected larval cadavers from different sources and geographic areas. The RAPD banding pattern of cadavers infected with A. aggregata differed from that of healthy leafcutting bee prepupae. RAPD analyses of cadavers infected with A. aggregata and A. larvis resulted in similar banding profiles as those obtained from corresponding pure fungal cultures of the two species. This suggests that the RAPD bands of infected cadavers were amplified from fungal DNA, rather than from other DNA associated with the leafcutting bee cadaver. The banding patterns of "sporulating" and "non-sporulating" chalkbrood cadavers exhibited no differences; this provides the first definitive evidence that both forms of the disease result from infection with A. aggregata.

Animals↗

PubMed1993

Localization and biochemical characterization of alfalfa mosaic virus replication complexes.

Replication complexes were isolated from alfalfa mosaic virus-infected tobacco protoplasts. Most of the RNA-synthesizing activity appears to colocalize with the intact chloroplasts upon sucrose-gradient centrifugation of cellular homogenates. Further analysis of these replication complexes showed that the enzyme is strongly associated with the outside of the chloroplasts, the endogenous template being well protected against ribonuclease action. RNA polymerase activity is sensitive to protease treatment of intact chloroplast fraction showing that an essential part of the enzyme complex is facing the in vitro medium, and probably the cytosol in vivo.

Cell Compartmentation↗

PubMed1993

Infection of tobacco with alfalfa mosaic virus cDNAs sheds light on the early function of the coat protein.

Coat protein (CP) is required in the inoculum to initiate infection of plants with the three genomic RNAs of alfalfa mosaic virus. Inoculation of plants with DNA copies of RNAs 1, 2, and 3, fused to the 35S promoter, resulted in virus replication but the infection level was increased several-fold by addition of CP to the inoculum. When one of the three cDNAs was replaced by its corresponding RNA molecule there was no infection unless CP was present in the inoculum. Plants transformed with a DNA copy of RNA 1 or RNA 2 could be infected with mixtures of cDNAs 2 and 3 or cDNAs 1 and 3, respectively. Again, when one cDNA in the inoculum was replaced by its corresponding RNA, infection depended on the presence of CP in the inoculum. However, when DNA copies of both RNA 1 and RNA 2 were present in the plant genome, the plants became infected after inoculation with cDNA 3 or RNA 3 without any requirement for CP. It is concluded that the necessity of CP in the inoculum depends on the timing of the production of the replicase subunits P1 and P2, encoded by RNAs 1 and 2, respectively, and on the availability of viral template RNAs once the replication complex has been formed. Models to explain the early function of CP are discussed.

Capsid↗

PubMed1993

Stress responses in alfalfa (Medicago sativa L.). 15. Characterization and expression patterns of members of a subset of the chalcone synthase multigene family.

We have identified five different full length chalcone synthase (CHS) cDNA clones from a cDNA library produced from transcripts isolated from an elicitor-treated alfalfa cell suspension culture. Nucleotide sequence similarity between the clones varied from 88-93%. Oligonucleotides based on divergent sequences in the 5'-untranslated regions of the clones could distinguish individual genes, or groups of genes, and their corresponding transcripts. Developmentally regulated expression of the CHS transcripts was predominantly in roots and root nodules; other unidentified members of the CHS gene family are expressed in stems, leaves and nodules. One of the CHS transcripts was strongly expressed in floral tissue. All the CHS transcripts studied were induced in elicitor-treated cell suspension cultures. Transcripts were also induced in roots in response to wounding or spraying with various elicitors, and in leaves infected with Phoma medicaginis (but not in wounded leaves). The induction kinetics of CHS2 transcripts were more rapid and/or transient than those of other members of the CHS family in CuCl2-treated roots and Phoma-infected leaves. The results are discussed in terms of the evolution and functions of the CHS gene family in legumes.

Acyltransferases↗

PubMed1994

Isolation of chalcone synthase and chalcone isomerase cDNAs from alfalfa (Medicago sativa L.): highest transcript levels occur in young roots and root tips.

Flavonoids are involved in several different interactions between plants and microorganisms. In the Rhizobium-legume symbiosis, they play an important role as inducers of rhizobial nodulation (nod) genes. We have identified from an alfalfa cDNA library four clones for chalcone synthase (CHS) and two clones for chalcone isomerase (CHI); CHS and CHI are key enzymes in flavonoid biosynthesis. In Medicago sp., CHS is encoded by 8-12 genes, and CHI is encoded by 1-2 genes. Here we report the DNA sequence of these clones as well as their relatedness to other legume CHS and CHI clones. In addition, we report on the expression patterns of two CHS gene family members as well as the CHI gene in M. sativa cv. Iroquois. While CHS and CHI transcript levels are high in root tips and entire young roots, they are low in effective nodules elicited by wild-type strains of Rhizobium meliloti and very low in aerial portions of the plant (stems, leaves, flowers). However, wounding the cotyledons results in a rapid increase in transcript levels of both chalcone synthase and chalcone isomerase genes in these organs.

Acyltransferases↗