| molecular cloning of dna complementary to tobacco vein mottling virus rna. | rna isolated from tobacco vein mottling virus (tvmv) was used as a template for avian myeloblastosis virus rna-dependent dna polymerase, primed with oligo(dt). the largest single-stranded cdna synthesized was 10 kb, the same as the viral rna. this material was converted to double-stranded cdna with escherichia coli dna polymerase i and digested with restriction endonuclease hindiii. the cdna fragments were ligated to hindiii-digested plasmid pbr322 and the product used to transform e. coli strai ... | 1983 | 18639027 |
| expression of alfalfa mosaic virus rna 4 cdna transcripts in vitro and in vivo. | copies of alfalfa mosaic virus (amv) rna 4 synthesized by transcription in vitro were shown to be biologically active in protoplasts when inoculated together with a mixture of amv rnas 1, 2, and 3. a 5' cap was essential for activity, whereas 15 nonviral nucleotides at the 5' end of the rna transcripts did not detectably affect activity. transcripts that terminated 65 nucleotides from the 3' end of the coding region and transcripts that lacked all or part of the 3'-noncoding region were effectiv ... | 1985 | 18640556 |
| amino acid preferences of retroviral proteases for amino-terminal positions in a type 1 cleavage site. | the specificities of the proteases of 11 retroviruses were studied using a series of oligopeptides with amino acid substitutions in the p1, p3, and p4 positions of a naturally occurring type 1 cleavage site (val-ser-gln-asn-tyr downward arrowpro-ile-val-gln) in human immunodeficiency virus type 1 (hiv-1). previously, the substrate specificity of the p2 site was studied for the same representative set of retroviral proteases, which included at least one member from each of the seven genera of the ... | 2008 | 18701588 |
| fbxw7 acts as an e3 ubiquitin ligase that targets c-myb for nemo-like kinase (nlk)-induced degradation. | the c-myb proto-oncogene product (c-myb) is degraded in response to wnt-1 signaling via a pathway involving tak1 (transforming growth factor-beta-activated kinase 1), hipk2 (homeodomain-interacting protein kinase 2), and nlk (nemo-like kinase). nlk directly binds to c-myb, which results in the phosphorylation of c-myb at multiple sites, and induces its ubiquitination and proteasome-dependent degradation. here, we report that fbxw7, the f-box protein of an scf complex, targets c-myb for degradati ... | 2008 | 18765672 |
| the use of elisa for detection of exogenous and endogenous avian leukosis viral antigens in basic breeding flocks. | an enzyme-linked immunosorbent assay (elisa) for avian leukosis/ sarcoma virus (alv) group-specific (gs) antigens was used to study the identification of hens which congenitally excrete exogenous alv. the sensitivity of this assay was compared with that of the phenotypic mixing test (pmt) and the direct complement fixation test (cft) by testing limiting dilutions of purified avian myeloblastosis virus (amv), embryo homogenates and albumens. about 0.4 ng/ml of purified amv protein could be detect ... | 1983 | 18766804 |
| application of monoclonal antibodies in the avian leukosis virus gs-antigen elisa. | five hybridoma cell lines which secrete antibodies to avian leukosis/ sarcoma (alv) group-specific (gs) antigens (gag gene products) have been established. the hybrid cells resulted from fusion of p3/x63-ag8.653 myeloma cells with splenocytes of balb/c mice which had been immunised with purified avian myeloblastosis virus (amv). screening of supernatant fluids was performed by an indirect double antibody sandwich enzyme-linked immunosorbent assay (idas-elisa) and the immunoelectroblotting techni ... | 1985 | 18766897 |
| detection of avian leukosis virus with the elisa system: evaluation of conjugation methodology and comparison of sensitivity with the phenotypic mixing test in commercial layer flocks. | conjugates of horseradish peroxidase with rabbit igg antibody against gs antigen (p27) of avian myeloblastosis virus were prepared using glutaraldehyde and periodate methods of coupling. conjugates were evaluated for the detection of gs antigen of avian leukosis and sarcoma viruses in the elisa system and the periodate conjugate was found to be superior. with an elisa based on a periodate conjugate it was possible to detect 5 x 10(3) iu of leukosis virus propagated in cell culture. comparisons b ... | 1982 | 18770224 |
| translation inhibition of capped and uncapped viral rnas mediated by ribosome-inactivating proteins. | abstract ribosome-inactivating proteins (rips) are n-glycosidases that remove specific purine residues from the sarcin/ricin (s/r) loop of the large rrna and arrest protein synthesis at the translocation step. in addition to their enzymatic activity, rips have been reputed to be potent antiviral agents against many plant, animal, and human viruses. we recently showed that pokeweed antiviral protein (pap), an rip from pokeweed, inhibits translation in cell extracts by binding to the cap structure ... | 2003 | 18942981 |
| identification, characterization, and molecular detection of alfalfa mosaic virus in potato. | abstract alfalfa mosaic virus (amv) was detected in potato fields in several provinces in canada and characterized by bioassay, enzyme-linked immunosorbent assay, and reverse-transcription polymerase chain reaction (rt-pcr). the identity of eight canadian potato amv isolates was confirmed by sequence analysis of their coat protein (cp) gene. sequence and phylogenetic analysis indicated that these eight amv potato isolates fell into one strain group, whereas a slight difference between ca175 and ... | 2006 | 18943961 |
| rna-dependent dna polymerases. | reverse transcriptases (rts) are multifunctional enzymes, but are mainly used as rna-directed dna polymerases in first-strand cdna synthesis. specifically, oligodeoxynucleotides are used as primers for extension on rna templates. the dna synthesized from an rna template is referred to as complementary dna (cdna) and is often used as a template for pcr or converted to dsdna for cloning. this unit describes appropriate reaction conditions for rts from moloney murine leukemia virus (mmlv) and avian ... | 2008 | 18972389 |
| characterization of moloney murine leukaemia virus/avian myeloblastosis virus chimeric reverse transcriptases. | reverse transcriptases (rts) from moloney murine leukaemia virus (mmlv) and avian myeloblastosis virus (amv) contain all the fingers, palm, thumb, connection and rnase h domains. the fingers, palm and thumb domains are thought to be involved in the reverse transcription activity, and the rnase h domain is in the rnase h activity. in this study, we characterized four chimeric rts which comprise one of the fingers, palm, thumb and rnase h domains originated from amv rt and the other three and conn ... | 2009 | 19060310 |
| transient protein expression in three pisum sativum (green pea) varieties. | the expression of proteins in plants both transiently and via permanently transformed lines has been demonstrated by a number of groups. transient plant expression systems, due to high expression levels and speed of production, show greater promise for the manufacturing of biopharmaceuticals when compared to permanent transformants. expression vectors based on a tobacco mosaic virus (tmv) are the most commonly utilized and the primary plant used, nicotiana benthamiana, has demonstrated the abili ... | 2009 | 19156736 |
| evaluation of different rt enzyme standards for quantitation of retroviruses using the single-tube fluorescent product-enhanced reverse transcriptase assay. | pcr-based reverse transcriptase (rt) assays are highly sensitive for broad detection of retroviruses. these assays are currently used for demonstrating the absence of retroviral contamination in vaccines and can also be applied to clinical and laboratory research to investigate low-virus replication. a single-tube fluorescent product-enhanced reverse transcriptase assay (stf-pert) has been published that was highly sensitive for retrovirus detection (<10 virions), with enhanced reproducibility a ... | 2009 | 19186191 |
| explorations into neolignan biosynthesis: concise total syntheses of helicterin b, helisorin, and helisterculin a from a common intermediate. | helicterins a and b (1 and 2), helisorin (3), and helisterculin a (4) are structurally unique natural products with the ability to combat the avian myeloblastosis virus. biogenetically, their architectures are considered to be products of seemingly straightforward diels-alder, radical-based, or acid-induced dimerizations of common, simpler precursors. yet, the pursuit of such blueprints in the laboratory has failed thus far in enabling their successful synthesis. herein, we describe the first to ... | 2009 | 19191699 |
| evidence for variation in the optimal translation initiation complex: plant eif4b, eif4f, and eif(iso)4f differentially promote translation of mrnas. | eukaryotic initiation factor (eif) 4b is known to interact with multiple initiation factors, mrna, rrna, and poly(a) binding protein (pabp). to gain a better understanding of the function of eif4b, the two isoforms from arabidopsis (arabidopsis thaliana) were expressed and analyzed using biophysical and biochemical methods. plant eif4b was found by ultracentrifugation and light scattering analysis to most likely be a monomer with an extended structure. an extended structure would facilitate the ... | 2009 | 19493973 |
| higher random oligo concentration improves reverse transcription yield of cdna from bioptic tissues and quantitative rt-pcr reliability. | real time quantitative reverse transcription-pcr (qrt-pcr) is the most sensitive technique for detection and quantification of mrna targets. reliable quantification of gene expression in formalin-fixed, paraffin-embedded tissues (ffpe), however, has been subjected to serious limitations so far, mainly due to the fragmentation of rna transcripts. we tried to improve the sensitivity and reliability of mrna quantification in ffpe by boosting the reverse transcription (rt) step, that is neglected in ... | 2009 | 19619529 |
| myb-induced chromatin remodeling at a dual enhancer/promoter element involves non-coding rna transcription and is disrupted by oncogenic mutations of v-myb. | the oncogene v-myb of avian myeloblastosis virus (amv) encodes a transcription factor (v-myb) that transforms myelomonocytic cells by deregulating the expression of specific target genes. v-myb has acquired its oncogenic potential by truncation as well as by a number of point mutations of its cellular progenitor c-myb. as a result of these changes, the target gene spectrum v-myb differs from that of c-myb. we recently showed that the chicken mim-1 gene, a c-myb target gene that is not activated ... | 2009 | 19841477 |
| in vitro and in vivo studies of the rna conformational switch in alfalfa mosaic virus. | the 3' termini of alfalfa mosaic virus (amv) rnas adopt two mutually exclusive conformations, a coat protein binding (cpb) and a trna-like (tl) conformer, which consist of a linear array of stem-loop structures and a pseudoknot structure, respectively. previously, switching between cpb and tl conformers has been proposed as a mechanism to regulate the competing processes of translation and replication of the viral rna (r. c. l. olsthoorn et al., embo j. 18:4856-4864, 1999). in the present study, ... | 2010 | 19923185 |
| complete genome amplification of equine influenza virus subtype 2. | this work reports a method for rapid amplification of the complete genome of equine influenza virus subtype 2 (h3n8). a thermoscript reverse transcriptase instead of the avian myeloblastosis virus reverse transcriptase or moloney murine leukemia virus reverse transcriptase was used. this enzyme has demonstrated higher thermal stability and is described as suitable to make long cdna with a complex secondary structure. the product obtained by this method can be cloned, used in later sequencing rea ... | 2009 | 20085182 |
| polymerase incorporation and miscoding properties of 5-chlorouracil. | inflammation-mediated hypochlorous acid (hocl) can damage dna, dna precursors, and other biological molecules, thereby producing an array of damage products such as 5-chlorouracil (clu). in this study, we prepared and studied 5-chloro-2'-deoxyuridine (cldu) and clu-containing oligonucleotide templates. we demonstrate that human k-562 cells grown in culture with 10 mum cldu incorporate substantial amounts of cldu without significant toxicity. when in the template, clu residues pair with datp but ... | 2010 | 20104909 |
| caulimoviridae tubule-guided transport is dictated by movement protein properties. | plant viruses move through plasmodesmata (pd) either as nucleoprotein complexes (npcs) or as tubule-guided encapsidated particles with the help of movement proteins (mps). to explore how and why mps specialize in one mechanism or the other, we tested the exchangeability of mps encoded by dna and rna virus genomes by means of an engineered alfalfa mosaic virus (amv) system. we show that caulimoviridae (dna genome virus) mps are competent for rna virus particle transport but are unable to mediate ... | 2010 | 20130061 |
| duplex structural differences and not 2'-hydroxyls explain the more stable binding of hiv-reverse transcriptase to rna-dna versus dna-dna. | human immunodeficiency virus reverse transcriptase (hiv-rt) binds more stably in binary complexes with rna-dna versus dna-dna. current results indicate that only the -2 and -4 rna nucleotides (-1 hybridized to the 3' recessed dna base) are required for stable binding to rna-dna, and even a single rna nucleotide conferred significantly greater stability than dna-dna. replacing 2'- hydroxyls on pivotal rna bases with 2'-o-methyls did not affect stability, indicating that interactions between hydro ... | 2010 | 20338878 |
| optimisation of reverse transcription can improve the sensitivity of rt-pcr for detection of classical swine fever virus. | classical swine fever is a highly contagious, notifiable disease of pigs and wild boars listed by the world organisation for animal health (oie). therefore, methods employed in the diagnosis of csf should be fast, sensitive and specific. the aim of this study was optimisation of the reverse transcription reaction to increase the sensitivity of real-time rt-pcr for the detection of classical swine fever virus, the aetiological agent of the disease. the efficiency of reverse transcription reaction ... | 2010 | 20460224 |
| alfalfa mosaic virus: coat protein-dependent initiation of infection. | summary taxonomy: alfalfa mosaic virus (amv) is the type species of the genus alfamovirus and belongs to the family bromoviridae. in this family, the tripartite rna genomes of bromo-, cucumo- and probably oleaviruses are infectious as such, whereas infection with the three genomic rnas of alfamo- and ilarviruses requires addition to the inoculum of a few molecules of coat protein (cp) per rna molecule. rnas 1 and 2 encode the replicase proteins p1 and p2, rna 3 encodes the movement protein and c ... | 2003 | 20569357 |
| effects of organic solvents on the reverse transcription reaction catalyzed by reverse transcriptases from avian myeloblastosis virus and moloney murine leukemia virus. | the use of certain organic chemicals has been found to improve yields and specificity in pcr. in this study, we examined the effects of dimethyl sulfoxide (dmso), formamide, and glycerol on the reverse transcription reaction catalyzed by reverse transcriptases (rts) from avian myeloblastosis virus (amv) and moloney murine leukemia virus (mmlv). at 42 °c, dmso at 24% v/v and formamide at 12-14% inhibited the cdna synthesis reaction, but dmso at 12% and formamide at 6-8% improved the efficiency of ... | 2010 | 20834159 |
| varied molecular interactions at the active sites of several dna polymerases: nonpolar nucleoside isosteres as probes. | we describe a survey of protein-dna interactions with seven different dna polymerases and reverse transcriptases, carried out with nonpolar nucleoside isosteres f (a thymidine analog) and z and q (deoxyadenosine analogues). previous results have shown that z and f can be efficiently replicated opposite each other by the exonuclease-free klenow fragment of dna polymerase i from escherichia coli (kf(-)), although both of them lack watson-crick h-bonding ability. we find that exonuclease-inactive t ... | 2000 | 20882113 |
| complete nucleotide sequence of a spanish isolate of alfalfa mosaic virus: evidence for additional genetic variability. | alfalfa mosaic virus (amv) is a plant virus that is distributed worldwide and can induce necrosis and/or yellow mosaic on a large variety of plant species, including commercially important crops. it is the only virus of the genus alfamovirus in the family bromoviridae. amv isolates can be clustered into two genetic groups that correlate with their geographic origin. here, we report for the first time the complete nucleotide sequence of a spanish isolate of amv found infecting cape honeysuckle (t ... | 2011 | 21327783 |
| c-myb in smooth muscle cells. | the proto-oncogene c-myb is a cellular homolog of a viral oncogene v-myb found in two independently derived avian acute leukemia viruses: avian myeloblastosis vn-us (amv) and e26 leukemia virus (1). the myb gene is highly conserved in eukaryotes, and it usually consists of 15 exons spanning over 35 kb of genomic dna (1).in humans, c-myb gene locus has been mapped to chromosome 6 (6q22-23) however, myb mrna expressed in thymus contains transcripts originating from chromosome 17 (17q25), suggestin ... | 1996 | 21359720 |
| synthesis of double-stranded complementary dna from poly(a)(+)mrna. | the use of avian myeloblastosis virus reverse transcriptase (amv rtase) to produce dna copies of mrna templates is a common and well-documented method (1-3). briefly, the method involves synthesis of a complementary dna strand to the mrna from a short double-stranded region, usually provided by using an oligo(dt) primer on poly(a)(+)rna. the enzyme does not always produce full length transcripts, but all the complementary strands are finished off with a short hairpin loop. this provides a ready- ... | 1985 | 21374188 |
| myb proteins talking to their dna (review). | dna sequence-specific proteins called transcription factors found in all multicellular organisms control the expression of genes and are involved in the regulation of the cell cycle. myb oncoproteins are transcription factors with a distinct dna binding domain lacking zinc fingers or a basic region. found originally in the avian myeloblastosis virus (v-myb), then in the genome of chickens (c-myb), the dna binding domain of myb occurs in regulatory proteins from mammals including humans, plants, ... | 1994 | 21559564 |
| comparison of the thermal stabilities of the αβ heterodimer and the α subunit of avian myeloblastosis virus reverse transcriptase. | avian myeloblastosis virus reverse transcriptase (amv rt) is a heterodimer consisting of a 63-kda α subunit and a 95-kda β subunit. in this study, we explored the role of the interaction between the α and β subunits on amv rt stability. the recombinant amv rt α subunit was expressed in insect cells and purified. it exhibited lower thermal stability than the native amv rt αβ heterodimer. unlike the αβ heterodimer, the α subunit was not stabilized by template-primer. these results suggest that int ... | 2011 | 21821920 |
| Detection of Murine Leukemia Virus or Mouse DNA in Commercial RT-PCR Reagents and Human DNAs. | The xenotropic murine leukemia virus (MLV)-related viruses (XMRV) have been reported in persons with prostate cancer, chronic fatigue syndrome, and less frequently in blood donors. Polytropic MLVs have also been described in persons with CFS and blood donors. However, many studies have failed to confirm these findings, raising the possibility of contamination as a source of the positive results. One PCR reagent, Platinum Taq polymerase (pol) has been reported to contain mouse DNA that produces f ... | 2011 | 22205995 |
| Status of tobacco viruses in Serbia and molecular characterization of tomato spotted wilt virus isolates. | In a four-year survey to determine the presence and distribution of viruses in tobacco crops at 17 localities of the Vojvodina Province and Central Serbia, 380 samples were collected and analyzed by DAS-ELISA. Out of the seven viruses tested, tomato spotted wilt virus (TSWV), potato virus Y (PVY), tobacco mosaic virus (TMV), cucumber mosaic virus (CMV), and alfalfa mosaic virus (AMV) were detected in 37.9, 33.4, 28.7, 23.9, and 15.5% of the total tested samples, respectively. TSWV was the most ... | 2011 | 22149499 |
| mechanism of hiv reverse transcriptase inhibition by zinc: formation of a highly stable enzyme-(primer-template) complex with profoundly diminished catalytic activity. | several physiologically relevant cations including ca(2+), mn(2+), and zn(2+) have been shown to inhibit hiv reverse transcriptase (rt), presumably by competitively displacing one or more mg(2+) ions bound to rt. we analyzed the effects of zn(2+) on reverse transcription and compared them to ca(2+) and mn(2+). using nucleotide extension efficiency as a readout, zn(2+) showed significant inhibition of reactions with 2 mm mg(2+), even when present at only ∼5 μm. mn(2+) and ca(2+) were also inhibit ... | 2011 | 21953456 |
| Molecular breeding of transgenic white clover (Trifolium repens L.) with field resistance to Alfalfa mosaic virus through the expression of its coat protein gene. | Viral diseases, such as Alfalfa mosaic virus (AMV), cause significant reductions in the productivity and vegetative persistence of white clover plants in the field. Transgenic white clover plants ectopically expressing the viral coat protein gene encoded by the sub-genomic RNA4 of AMV were generated. Lines carrying a single copy of the transgene were analysed at the molecular, biochemical and phenotypic level under glasshouse and field conditions. Field resistance to AMV infection, as well as mi ... | 2011 | 21947755 |
| Rapid detection of wheat yellow mosaic virus by reverse transcription loop-mediated isothermal amplification. | ABSTRACT: For the detection of wheat yellow mosaic virus (WYMV), we established a reverse transcription loop-mediated isothermal amplification (RT-LAMP) method. Using Primer Explorer software, four sets of primers were designed and RT-LAMP assay reaction conditions were optimized. The RT-LAMP was performed at different times by four primer sets. Agarose gel analysis showed that WYMV could be detected after 30 min with the primer set III and after 45 min with the other three primer sets, both und ... | 2011 | 22185375 |
| microwave-assisted rapid enzymatic synthesis of nucleic acids. | herein we report microwave-induced enhancement of the reactions catalyzed by escherichia coli dna polymerase i and avian myeloblastosis virus-reverse transcriptase. the reactions induced by microwaves result in a highly selective synthesis of nucleic acids in 10-50 seconds. in contrast, same reactions failed to give desired reaction products when carried out in the same time periods, but without microwave irradiation. each of the reactions was carried out for different duration of microwave expo ... | 2016 | 27159147 |
| kl-6 regulated the expression of hgf, collagen and myofibroblast differentiation. | kl-6 is a pulmonary epithelial-derived mucin which is secreted mainly by type ii alveolar epithelial cells. the level of kl-6 in serum is closely correlated to the clinical activity of various interstitial lung diseases (ild) and acts as a prognostic factor for ild patients. previous studies have showed that kl-6 promoted chemotaxis, proliferation and inhibited the apoptosis of human lung fibroblasts. however, the underlying mechanism remains unknown. in this study, we investigated the function ... | 2013 | 24302189 |
| base-pairing preferences, physicochemical properties and mutational behaviour of the dna lesion 8-nitroguanine. | 8-nitro-2'-deoxyguanosine (8-nitrodg) is a relatively unstable, mutagenic lesion of dna that is increasingly believed to be associated with tissue inflammation. due to the lability of the glycosidic bond, 8-nitrodg cannot be incorporated into oligodeoxynucleotides (odns) by chemical dna synthesis and thus very little is known about its physicochemical properties and base-pairing preferences. here we describe the synthesis of 8-nitro-2'-o-methylguanosine, a ribonucleoside analogue of this lesion, ... | 2012 | 22965127 |
| viral reverse transcriptases show selective high affinity binding to dna-dna primer-templates that resemble the polypurine tract. | previous results using a selex (systematic evolution of ligands by exponential enrichment)-based approach that selected dna primer-template duplexes binding with high affinity to hiv reverse transcriptase (rt) showed that primers mimicking the 3' end, and in particular the six nt terminal g tract, of the rna polypurine tract (ppt; hiv ppt: 5'-aaaagaaaagggggg-3') were preferentially selected. in this report, two viral (moloney murine leukemia virus (mulv) and avian myeloblastosis virus (amv)) and ... | 2012 | 22848574 |
| improving the thermal stability of avian myeloblastosis virus reverse transcriptase α-subunit by site-directed mutagenesis. | avian myeloblastosis virus reverse transcriptase (amv rt) is a heterodimer consisting of a 63 kda α-subunit and a 95 kda β subunit. moloney murine leukaemia virus reverse transcriptase (mmlv rt) is a 75 kda monomer. these two rts are the most extensively used for conversion of rna to dna. we previously developed several mutations that increase the thermostability of mmlv rt and generated a highly stable mmlv rt variant e286r/e302k/l435r/d524a by combining three of them (glu286→arg, glu302→lys, a ... | 2012 | 22426840 |
| a colony hybridization study of mrna in maturating soybean seeds. | polysomal poly(a)rna was transcribed by avian myeloblastosis virus (amv) reverse transcriptase and double-stranded complementary dna (cdna) was then synthesized and cloned in escherichia coli c600 by (dg)-(dc) tailing in the psti site of pbr322. a bank of mrna sequences from maturating soybean seeds (average weight 200 mg) was obtained. (32)p-cdna based on mrna from various organs of the plant (leaves, axes, seeds) was synthesized. the clones were hybridized with different kinds of (32)p-cdna on ... | 1982 | 24257772 |