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protein expression in e. coli minicells by recombinant plasmids.the polypeptides synthesized in e. coli minicells from recombinant plasmids containing dna fragments from cauliflower mosaic virus, drosophila melanogaster, and mouse mitochondria were examined. molecularly cloned fragments of cauliflower mosaic virus dna directed the synthesis of high levels of three polypeptides, which were synthesized entirely from within the cloned virus dna fragments independent of their insertion into the plasmid vehicles. several fragments of d. melanogaster dna were capa ...1977403011
cloning of cauliflower mosaic virus (clmv) dna in escherichia coli.a plasmid containing cauliflower mosaic virus dna can be faithfully cloned in escherichia coli, but proved to be noninfective in test plants.1977322284
the structure of cauliflower mosaic virus. i. a restriction endonuclease map of cauliflower mosaic virus dna. 1977329557
the pff plasmids: cassettes utilising camv sequences for expression of foreign genes in plants.a plant expression cassette was constructed using the cauliflower mosaic virus 35s 5' regulatory region with the enhancer duplicated and the 35s polyadenylation signal. insertion of a polylinker between the transcription initiation and polyadenylation sites allows for easy cloning of genes. to test the usefulness of the cassette chimeric bacterial genes were prepared. the constructs were introduced into nicotiana tabacum suspension culture cells by the particle bombardment process. expression of ...19901369289
agroinfection as an alternative to insects for infecting plants with beet western yellows luteovirus.beet western yellows luteovirus, like other luteoviruses, cannot be transmitted to host plants by mechanical inoculation but requires an aphid vector, a feature that has heretofore presented a serious obstacle to the study of such viruses. in this paper we describe use of agroinfection to infect hosts with beet western yellows virus without recourse to aphids. agroinfection is a procedure for introducing a plant virus into a host via agrobacterium tumefaciens harboring a ti plasmid, which can ef ...19921409615
effect of two consensus sequences preceding the translation initiator codon on gene expression in plant protoplasts.expression cassettes containing a duplicated cauliflower mosaic virus (camv) 35s promoter fused to a polylinker preceded by the ccaccatgg and aacaatgg sequences were constructed. these two sequences correspond to the consensus sequences around the translation start codons in vertebrates and plants respectively. translational fusions were made with the beta-glucuronidase-coding sequence and transient expression was recorded in tobacco mesophyll protoplasts. approximately three times more gus acti ...19921373083
photoreactivation of dna-containing cauliflower mosaic virus and tobacco mosaic virus rna on datura. 1977905348
metabolic engineering of medicinal plants: transgenic atropa belladonna with an improved alkaloid composition.the tropane alkaloid scopolamine is a medicinally important anticholinergic drug present in several solanaceous plants. hyoscyamine 6 beta-hydroxylase (ec 1.14.11.11) catalyzes the oxidative reactions in the biosynthetic pathway leading from hyoscyamine to scopolamine. we introduced the hydroxylase gene from hyoscyamus niger under the control of the cauliflower mosaic virus 35s promoter into hyoscyamine-rich atropa belladonna by the use of an agrobacterium-mediated transformation system. a trans ...19921465402
different sequence elements are required for function of the cauliflower mosaic virus polyadenylation site in saccharomyces cerevisiae compared with in plants.we show that the polyadenylation site derived from the plant cauliflower mosaic virus (camv) is specifically functional in the yeast saccharomyces cerevisiae. the mrna 3' endpoints were mapped at the same position in yeast cells as in plants, and the camv polyadenylation site was recognized in an orientation-dependent manner. mutational analysis of the camv 3'-end-formation signal revealed that multiple elements are essential for proper activity in yeast cells, including two upstream elements th ...19921373813
definition of the upstream efficiency element of the simian virus 40 late polyadenylation signal by using in vitro analyses.the polyadenylation signal for the late mrnas of simian virus 40 is known to have sequence elements located both upstream and downstream of the aauaaa which affect efficiency of utilization of the signal. the upstream efficiency element has been previously characterized by using deletion mutations and transfection analyses. those studies suggested that the upstream element lies between 13 and 48 nucleotides upstream of the aauaaa. we have utilized in vitro cleavage and polyadenylation reactions ...19921333042
two g-box-related sequences confer different expression patterns in transgenic tobacco.we have analyzed the expression patterns conferred by two g-box-related motifs, a perfect palindromic sequence (pa, 5'-gccacgtggc-3') and motif i (iwt, 5'-gtacgtggcg-3'), in transgenic tobacco plants. a mutant version of motif i, imu, was used as a negative control. pa is present in the promoters of several different genes, whereas iwt is a conserved sequence found in abscisic acid-inducible promoters. previously we have demonstrated that pa and iwt, but not imu, can bind to the tobacco transcri ...19921467649
effective vectors for transformation, expression of heterologous genes, and assaying transposon excision in transgenic plants.progress in plant molecular biology has depended heavily on the availability of effective vectors for plant cell transformation and heterologous expression. in this paper we describe the structures of a wide array of plasmids which have proved extremely effective in (a) plant transformation, (b) expression of heterologous genes and (c) assaying the activity of transposons in transgenic plants. constructs that confer resistance to kanamycin, hygromycin, streptomycin, spectinomycin and phosphinotr ...19921338696
the 5'-untranslated leader sequence of potato virus x rna enhances the expression of a heterologous gene in vivo.the 5' untranslated leader of potato virus x (pvx) rna is shown when contiguous to the coding sequence, to enhance the expression of the neomycin phosphotransferase ii gene (nptii) in nicotiana tabacum protoplasts in vivo. the level of transient expression of the nptii gene in protoplasts provided by a plasmid containing the coding sequence of the nptii gene under the control of 35s cauliflower mosaic virus (camv) promoter and terminator elements served as the baseline control. insertion of the ...19921324400
high rates of ac/ds germinal transposition in arabidopsis suitable for gene isolation by insertional mutagenesis.overexpression of the activator (ac) transposase gene in arabidopsis thaliana resulted in a minimal germinal transposition frequency of 27% in which independent dissociation (ds) transposition events were observed. molecular analysis of 45 f1 generation ac/ds plants indicated that high rates of somatic excision had occurred, and independent germinal insertions were identified in f2 generation progeny plants. a tandem cauliflower mosaic virus (camv) promoter fused to two different ac coding seque ...19921321434
stringent repression and homogeneous de-repression by tetracycline of a modified camv 35s promoter in intact transgenic tobacco plants.a cauliflower mosaic virus (camv) 35s promoter derivative, which is tightly repressed by the tn 10 encoded tet repressor in a transient expression system as well as in transgenic plants has been constructed. after treatment of transgenic plants with tetracycline (tc) the activity of the reporter enzyme beta-glucuronidase (gus) increased up to 500-fold in tissue culture as well as under greenhouse conditions. efficient de-repression was achieved by tc uptake through the roots as well as by tc tre ...19921303802
expression in transgenic tobacco of the bacterial neomycin phosphotransferase gene modified by intron insertions of various sizes.a plant selectable marker gene consisting of cauliflower mosaic virus expression signals and the protein-coding sequence of bacterial neomycin phosphotransferase was modified by insertion of an intron sequence from a storage protein gene, phaseolin. correct and efficient splicing of the resulting mosaic rna was observed in transgenic tobacco plants. the insertion of various linkers or gradual increase of intron size by addition in both orientations of internal intron sequences from another plant ...19921322741
synthetic cryiiia gene from bacillus thuringiensis improved for high expression in plants.a 1974 bp synthetic gene was constructed from chemically synthesized oligonucleotides in order to improve transgenic protein expression of the cryiiia gene from bacillus thuringiensis var. tenebrionis in transgenic tobacco. the crystal toxin genes (cry) from b. thuringiensis are difficult to express in plants even when under the control of efficient plant regulatory sequences. we identified and eliminated five classes of sequence found throughout the cryiiia gene that mimic eukaryotic processing ...19921301214
a restriction map of cauliflower mosaic virus dna (strain pv 147). mapping of the cleavage sites of hhai, saci, avai, pvuii, psti, xbai, ecori, bg/ii, hincii, hpaii and hindii + iii.the virion-extracted dna (mr5 x 10(6)) of cauliflower mosaic virus (camv) has three single-stranded interruptions. the mapping of this dna using eleven restriction endonucleases (hhai, saci, avai, pvuii, psti, xbai, ecori, bg/ii, hincii, hpaii and hindii + iii) is reported here. the existence of the three single-stranded breaks complicates the identification and the molecular weight determination of fragments produced by hpaii, hindiii and hindii + iii. indeed the electrophoretic mobility of som ...1979488094
expression of cauliflower mosaic virus orf ii in a baculovirus system.the cauliflower mosaic virus orf ii encoding the aphid transmission factor (atf) was mutagenized to introduce a bamhi restriction site upstream from the initiation codon and then cloned into an eukaryotic viral expression vector (autographa californica nuclear polyhedrosis virus). all recombinant viruses tested in spodoptera frugiperda (sf21) cells expressed a protein of about 18 kd which comigrated in page with atf from infected plants. western blotting using an oligopeptide antiserum to atf co ...19921428752
expression of a coriander desaturase results in petroselinic acid production in transgenic tobacco.little is known about the metabolic origin of petroselinic acid (18:1 delta 6cis), the principal fatty acid of the seed oil of most umbelliferae, araliaceae, and garryaceae species. to examine the possibility that petroselinic acid is the product of an acyl-acyl carrier protein (acp) desaturase, western blots of coriander and other umbelliferae seed extracts were probed with antibodies against the delta 9-stearoyl-acp desaturase of avocado. in these extracts, proteins of 39 and 36 kda were detec ...19921454797
maize polyubiquitin genes: structure, thermal perturbation of expression and transcript splicing, and promoter activity following transfer to protoplasts by electroporation.two genomic clones (lambda ubi-1 and lambda ubi-2) encoding the highly conserved 76 amino acid protein ubiquitin have been isolated from maize. sequence analysis shows that both genes contain seven contiguous direct repeats of the protein coding region in a polyprotein conformation. the deduced amino acid sequence of all 14 repeats is identical and is the same as for other plant ubiquitins. the use of transcript-specific oligonucleotide probes shows that ubi-1 and ubi-2 are expressed constitutiv ...19921313711
biologically active cymbidium ringspot virus satellite rna in transgenic plants suppresses accumulation of di rna.a full-length dna copy of cymbidium ringspot virus (cyrsv) satellite rna was cloned downstream of the bacteriophage t7 rna polymerase promoter. in vitro transcripts were biologically active in plants when coinoculated with the helper virus or its rna. although the transcripts contained 7 or 29 extra nucleotides at the 3' end, the proper 3' terminus was restored in the satrna progeny. full-length cdna clones of cyrsv satrna under the control of the cauliflower mosaic virus 35s promoter and termin ...19921374981
arabidopsis thaliana small subunit leader and transit peptide enhance the expression of bacillus thuringiensis proteins in transgenic plants.the expression of the modified gene for a truncated form of the cryia(c) gene, encoding the insecticidal portion of the lepidopteran-active cryia(c) protein from bacillus thuringiensis var. kurstaki (b.t.k.) hd73, under control of the arabidopsis thaliana ribulose-1,5-bisphosphate carboxylase (rubisco) small subunit ats1a promoter with and without its associated transit peptide was analyzed in transgenic tobacco plants. examination of leaf tissue revealed that the ats1a promoter with its transit ...19921515613
expression of a viral avirulence gene in transgenic plants is sufficient to induce the hypersensitive defense reaction.tobacco plants containing the n' resistance gene exhibit a hypersensitive defense reaction when infected with tomato mosaic virus (tomv); infection results in necrotic lesions at the primary infection sites. in an attempt to investigate the molecular mechanism(s) underlying this plant-pathogen interaction, the tomv coat protein gene was joined by a transcriptional fusion to the strong constitutive 35s rna promoter from cauliflower mosaic virus. this chimeric gene was introduced via agrobacterium ...19921515667
localization of cauliflower mosaic virus in the cell nucleus of brassica pekinensis l.cauliflower mosaic virus (camv) particles were observed in the nuclei of xylem parenchyma cells in brassica pekinensis l. doubly infected by camv and turnip mosaic virus (tumv). camv particles were aggregated in the nucleoplasm but not embedded in viroplasms. this phenomenon was not detected in cell nuclei of mesophyll tissue. typical features associated with infection by either camv or tumv normally occurred in the cytoplasm of cells of both tissues: two types of viroplasms with embedded camv p ...19921410828
development and characterization of a generalized gene tagging system for higher plants using an engineered maize transposon ac.this report describes a series of transposon tagging vectors for dicotyledonous plants based on the maize transposable element ac. this binary system includes the transposase (ts) and the tagging element (ds) on separate t-dna vectors. ts elements include versions in which transcription is driven either by the endogenous ac promoter or by the cauliflower mosaic virus (camv) 35s promoter. ds tagging element includes a gene conferring methotrexate (mtx) resistance for selection and a supf gene to ...19921327269
expression of a calmodulin methylation mutant affects the growth and development of transgenic tobacco plants.transgenic plants were constructed that express two foreign calmodulins (vu-1 and vu-3 calmodulins) derived from a cloned synthetic calmodulin gene. vu-1 calmodulin, similar to endogenous plant calmodulin, possesses a lysine residue at position 115 and undergoes posttranslational methylation. vu-3 calmodulin is a site-directed mutant of vu-1 calmodulin that is identical in sequence except for the substitution of an arginine at position 115 and thus is incapable of methylation. both calmodulin ge ...19921325656
the cauliflower mosaic virus 35s promoter is regulated by camp in saccharomyces cerevisiae.the cauliflower mosaic virus 35s promoter confers strong gene expression in plants, animals and fission yeast, but not in budding yeast. on investigating this paradox, we found that in budding yeast the promoter acts through two domains. whereas the upstream domain acts as a silencer, the downstream domain couples expression to the nutritional state of the cells via the ras/camp pathway. point mutations indicate that two boxes with similarity to the camp regulated element (cre) of mammalian cell ...19921334531
cauliflower mosaic virus reverse transcriptase. activation by proteolytic processing and functional alteration by terminal deletion.we have previously expressed the cauliflower mosaic virus (camv) reverse transcriptase (rtase) gene, the orfv gene, in yeast in an active form (rtase-y). an activity gel analysis revealed that the molecular size of rtase-y as well as an rtase associated with the camv particles (rtase-v) is 60 kda. this size is about 18 kda smaller than that of the inactive form previously expressed in escherichia coli (rtase-e) (78 kda), which corresponds to the coding capacity estimated for the orfv gene. to in ...19921375943
role of an upstream open reading frame in the translation of polycistronic mrnas in plant cells.the influence of an upstream small open reading frame (urf) on the translation of two consecutive coding regions on an eukaryotic mrna was studied. the cis effects of leader length, urf length, the sequences of the urf and neighboring regions, and the trans effects of the cauliflower mosaic virus transactivator (tav) were analyzed. translation efficiency of the immediate downstream open reading frame (orf) decreased with increasing urf length. short urfs did not drastically inhibit translation o ...19921508670
characterization of cymbidium mosaic virus coat protein gene and its expression in transgenic tobacco plants.cymbidium mosaic virus (cymv) is the most prevalent virus infecting orchids. here, we report the isolation of partial cdna clones encoding the genomic rna of cymv. like most of the polyadenylated monopartite positive-strand rna viruses, the open reading frame (orf) coding for the viral coat protein (cp) is located at the 3' end. the orf predicts a polypeptide chain of 220 amino acids with a molecular weight of 23,600. sequence comparison of this orf to the cp sequences of potato virus x(pvx) and ...19921600145
promoter fusions to the activator transposase gene cause distinct patterns of dissociation excision in tobacco cotyledons.to explore the effects of altering the level of activator (ac) transposase (tpase) expression, a series of plasmids was constructed in which heterologous promoters were fused to the tpase gene. promoters for the cauliflower mosaic virus (camv) 35s transcript and the octopine synthase (ocs) and nopaline synthase (nos) genes were tested. these fusions, and constructs expressing tpase from the wild-type ac promoter, were introduced into tobacco, and their activity was monitored by crossing to a lin ...19921323365
elevated levels of activator transposase mrna are associated with high frequencies of dissociation excision in arabidopsis.the activator (ac) element of maize is active at a low frequency in arabidopsis. to determine whether this is due to poor expression of the ac transposase gene, we obtained and studied 19 arabidopsis transformants containing fusions of the octopine synthase (ocs), nopaline synthase (nos), cauliflower mosaic virus (camv) 35s, or ac promoters to the transposase open reading frame. these transformants were examined both for their ability to drive excision of a dissociation (ds) element from a strep ...19921323366
control of gene expression in tobacco cells using a bacterial operator-repressor system.we have investigated the efficacy of using the escherichia coli lac operator-repressor system to control plant gene expression. the laci gene was modified to allow optimal expression in plant cells and then placed downstream of the cauliflower mosaic virus (camv) 35s rna promoter. this construct was introduced into tobacco plants by leaf disc transformation. transgenic tobacco plants synthesized significant quantities of laci protein (up to 0.06% of total soluble protein). we have used the e.col ...19921563343
biophysical and biochemical properties of baculovirus-expressed camv p1 protein.cauliflower mosaic virus (camv) gene i encodes a protein (p1) that has been implicated in the control of virus movement in infected plants. to assist in the characterization of the mechanism of action of p1, gene i has been expressed efficiently in spodoptera frugiperda (sf) cells using recombinant baculovirus. control of the expression of camv gene i by the polyhedrin late promoter in the baculovirus autographa californica nuclear polyhedrosis virus (acnpv) resulted in very high levels of p1 ac ...19921549910
use of the signal peptide of pisum vicilin to translocate beta-glucuronidase in nicotiana tabacum.a hybrid protein system was used for the study of protein transport in plant cells. a nucleotide sequence (vic) encoding a putative signal peptide of 15 amino acid residues, derived from the published aa sequence of one pisum vicilin, was synthesized and fused in frame to the gus gene encoding a bacterial cytosolic beta-glucuronidase (gus). when the hybrid vic::gus gene was expressed in tobacco cells using the cauliflower mosaic virus 35s promoter, the hybrid gus protein was targeted to, and gly ...19921555771
expression of cauliflower mosaic virus gene i in saccharomyces cerevisiae.cauliflower mosaic virus (camv) gene i encodes a 40-kda protein, p1, which is thought to be involved in the cell-to-cell movement of the virus. in order to investigate its functioning, p1 was expressed in saccharomyces cerevisiae transformed by an expression vector containing camv gene i. when produced in yeast, pi was 40 kda in size and not n-glycosylated.19911796216
regulated inactivation of homologous gene expression in transgenic nicotiana sylvestris plants containing a defense-related tobacco chitinase gene.the class i chitinases are vacuolar proteins implicated in the defense of plants against pathogens. leaves of transgenic nicotiana sylvestris plants homozygous for a chimeric tobacco (nicotiana tabacum) chitinase gene with cauliflower mosaic virus (camv) 35s rna expression signals usually accumulate high levels of chitinase relative to comparable leaves of non-transformed plants. unexpectedly, some transgenic plants accumulated lower levels of chitinase than nontransformed plants. we call this p ...19921281514
processing of the minor capsid protein of the cauliflower mosaic virus requires a cysteine proteinase.the major capsid protein of the cauliflower mosaic virus (camv) is processed in vivo. the viral aspartic proteinase that catalyses this maturation has been characterized previously and is coded by the camv gene v. this virus has a second capsid protein, a minor component, encoded by gene iii. this protein, p3, is also processed at its c-terminus in vivo. to determine whether p3 is matured by the camv proteinase p5, we expressed, in saccharomyces cerevisiae, p3, p5 and a fusion protein p7-p4, con ...19921480825
a versatile binary vector system with a t-dna organisational structure conducive to efficient integration of cloned dna into the plant genome.a versatile gene expression cartridge and binary vector system was constructed for use in agrobacterium-mediated plant transformation. the expression cartridge of the primary cloning vector, part7, comprises of cauliflower mosaic virus cabb b-ji isolate 35s promoter, a multiple cloning site and the transcriptional termination region of the octopine synthase gene. the entire cartridge can be removed from part7 as a not i fragment and introduced directly into the binary vector, part27, recombinant ...19921463857
mnf1, a leaf tissue-specific dna-binding protein of maize, interacts with the cauliflower mosaic virus 35s promoter as well as the c4 photosynthetic phosphoenolpyruvate carboxylase gene promoter.when gel shift assays were performed with maize nuclear extract and a dna fragment containing the cauliflower mosaic virus (camv) 35s promoter, three dna-protein complexes were observed. analyses with nuclear extracts prepared from green leaves, etiolated leaves, stems and roots showed that the complexes resulted from the existence of at least two nuclear factors. one of them is presumably a constitutive nuclear factor found in all tissues tested, and another is a leaf-specific factor present bo ...19921627769
suppression of gene expression in plant cells utilizing antisense sequences transcribed by rna polymerase iii.inverted sequences of the chloramphenicol acetyltransferase (cat) reporter gene were fused to a soybean trna(met(i)) gene lacking a terminator such that the trna(met(i)) sequences caused the co-transcription of cat antisense sequences by rna polymerase iii. when electroporated into carrot protoplasts, these antisense dna constructs suppressed cat enzyme activity expressed from co-electroporated dnas containing the cat gene downstream of the cauliflower mosaic virus (camv) 35s rna promoter. our m ...19921627777
transient expression of the coat protein of sugarcane mosaic virus in sugarcane protoplasts and expression in escherichia coli.the coat protein (cp) of strain sc of sugarcane mosaic virus (scmv-sc) was expressed transiently in sugarcane protoplasts after electroporation with one of two plasmids encoding the cp gene. the cp gene was fused with either the cauliflower mosaic virus 35s promoter or the synthetic monocotyledon promoter "emu". the coat protein gene was also inducibly expressed in escherichia coli when fused to the trc promoter. the protein expressed in both systems had the same electrophoretic mobility and ant ...19921642548
the function of vacuolar beta-1,3-glucanase investigated by antisense transformation. susceptibility of transgenic nicotiana sylvestris plants to cercospora nicotianae infection.vacuolar class i beta-1,3-glucanases (ec 3.2.1.39) are believed to be important in the induced defense reaction of plants to fungal infection. we used antisense transformation to test this hypothesis and to identify other possible physiological functions of this enzyme. nicotiana sylvestris plants were transformed with antisense constructions containing the region from position 27 to 608 of the coding sequence of the basic, vacuolar beta-1,3-glucanase gene gla of tobacco regulated by cauliflower ...19921643283
regulation of the activities of african cassava mosaic virus promoters by the ac1, ac2, and ac3 gene products.dna fragments comprising each of the promoter regions from the geminivirus african cassava mosaic virus (acmv) were cloned into the puc18-based vector, pg1, producing transcriptional fusions with the beta-glucuronidase gene (gus) and nopaline synthase terminator sequence. the relative activity of each promoter construct was analyzed by a gus expression assay of extracts from nicotiana clevelandii protoplasts coelectroporated with the gus reporter constructs and constructs in which individual acm ...19921585657
two proteins encoded by beet necrotic yellow vein virus rna 3 influence symptom phenotype on leaves.rna 3 of the beet necrotic yellow vein virus (bnyvv) quadripartite rna genome is not essential for virus multiplication on leaves of tetragonia expansa but has dramatic effects on symptom expression. virus isolates containing rna 3 produce bright yellow local lesions while isolates lacking rna 3 produce much milder symptoms. using directed mutagenesis of cdna clones followed by in vitro synthesis of biologically active transcripts, a 25 kda open reading frame (orf) of rna 3 was shown to be respo ...19921537331
effects of thionins on beta-glucuronidase in vitro and in plant protoplasts.thionins cause the irreversible inactivation of beta-glucuronidase (gus) in vitro in a dose- and time-dependent manner. the enzyme is also sensitive to externally added thionins when expressed in the cytoplasmic compartment of tobacco protoplasts transformed with the gus gene under the 35s promoter of the cauliflower mosaic virus. in protoplasts transformed with the gus gene fused to a signal peptide, where gus is translocated into the lumen of the endoplasmic reticulum, the activity is signific ...19921537404
quantification and comparison of chloramphenicol acetyltransferase activity in transformed plant protoplasts using high-performance liquid chromatography- and radioisotope-based assays.rice and petunia leaf and cell suspension protoplasts were transformed by electroporation in the presence of pdw2. this plasmid contains a chloramphenicol acetyltransferase (cat) coding region under the control of a promoter constructed from sequences of the cauliflower mosaic virus genome. we have compared two different approaches to measuring cat activity in this system, namely high-performance liquid chromatography (hplc) and a radioisotope-based method. our results show that both techniques ...19921621965
nucleic acid-binding properties of the alfalfa mosaic virus movement protein produced in yeast.the movement protein of alfalfa mosaic virus (p3) was purified from yeasts transformed with an expression vector containing the p3 gene. its nucleic acid-binding properties were tested by electrophoretic retardation, nitrocellulose retention, and rna-protein cross-linking. the recombinant protein had a higher affinity for single-stranded rna and dna than for double-stranded nucleic acids. each nucleic acid molecule bound several protein molecules without sequence specificity. the binding was 80% ...19921585656
agroinfection of transgenic plants leads to viable cauliflower mosaic virus by intermolecular recombination.intermolecular reconstitution of a plant virus has been detected in whole plants in a system using a defective cauliflower mosaic virus genome and transgenic host plants containing the missing viral gene. the information for the gene vi protein of the virus was integrated into the chromosome of host brassica napus plants and leaves of these plants were inoculated with agrobacterium tumefaciens containing the complementing viral sequences. in several cases, upper leaves contained replicating vira ...19921546451
a cdna clone of tomato mosaic virus is infectious in plants.a cdna clone of tomato mosaic virus (tomv) genomic rna was fused to the cauliflower mosaic virus 35s rna promoter and the nopaline synthase gene polyadenylation signal. the transcriptional initiation site of the 35s rna promoter was altered by in vitro mutagenesis so that the resulting transcripts start at the first nucleotide of the tomv sequence. in addition, 12 nucleotides were inserted in the 5' untranslated region of the tomv genome. this plasmid, psln, was used to inoculate several host pl ...19921583735
infectious in vivo transcripts of a plum pox potyvirus full-length cdna clone containing the cauliflower mosaic virus 35s rna promoter.a full-length cdna clone of an aphid non-transmissible isolate of plum pox potyvirus (ppv) was rendered biologically active when placed under the control of the cauliflower mosaic virus 35s rna promoter and the nopaline synthase polyadenylation signal. the cdna was constructed so that the exact 5' end of the ppv rna was present at the transcription initiation site. inoculation of plasmid dna onto nicotiana benthamiana led to systemic infection, whereas local lesions were produced in chenopodium ...19921545225
in vitro transcription from cauliflower mosaic virus promoters by a cell-free extract from tobacco cells.we have studied transcription from the cauliflower mosaic virus 19s and 35s promoters in a cell-free system derived from tobacco cells in suspension culture. while a whole-cell extract is incapable of detectable transcription from these promoters, successive purification by column chromatography allows the preparation of two fractions which contain all factors necessary for transcription from the 19s promoter. in contrast, transcription from the 35s promoter leads to the accumulation of short rn ...19901715207
the commelina yellow mottle virus promoter is a strong promoter in vascular and reproductive tissues.commelina yellow mottle virus (coymv) is a double-stranded dna virus that infects the monocot commelina diffusa. although coymv and cauliflower mosaic virus (camv; another double-stranded dna virus) probably replicate by a similar mechanism, the particle morphology and host range of coymv place it in a distinct group. we present evidence that a prompter fragment isolated from coymv confers a tissue-specific pattern of expression that is different from that conferred by the camv 35s promoter. whe ...19921633493
inhibition of the expression of the gene for granule-bound starch synthase in potato by antisense constructs.granule-bound starch synthase [gbss; ec 24.1.21] determines the presence of amylose in reserve starches. potato plants were transformed to produce antisense rna from a gene construct containing a full-length granule-bound starch synthase cdna in reverse orientation, fused between the cauliflower mosaic virus 35s promoter and the nopaline synthase terminator. the construct was integrated into the potato genome by agrobacterium rhizogenes-mediated transformation. inhibition of gbss activity in pot ...19912005870
pea lectin is correctly processed, stable and active in leaves of transgenic potato plants.a gene encoding the preproprotein of the pea (pisum sativum) lectin was expressed in transgenic potato plants using a cauliflower mosaic virus (camv) 35s promoter or a tobacco ribulose bisphosphate carboxylase small subunit (ssrubisco) promoter. presence of the pea lectin to levels greater than 1% of total soluble leaf protein was detected by radioimmunoassay (ria). the pattern of expression derived from the two promoters was established using both ria and a squash-blot immunolocalisation techni ...19911868225
the indoleacetic acid-lysine synthetase gene of pseudomonas syringae subsp. savastanoi induces developmental alterations in transgenic tobacco and potato plants.the iaal gene of pseudomonas syringae subsp. savastanoi encodes an indoleacetic acid-lysine synthetase that conjugates lysine to indoleacetic acid. a chimaeric gene consisting of the iaal coding region under the control of the 35s rna promoter from cauliflower mosaic virus (35siaal) has been used to test if iaal gene expression leads to morphological alterations in tobacco and potato. transgenic tobacco plantlets bearing this construct have been shown to synthesize iaa-[14c]lysine when fed with ...19911905782
properties of commelina yellow mottle virus's complete dna sequence, genomic discontinuities and transcript suggest that it is a pararetrovirus.the non-enveloped bacilliform viruses are the second group of plant viruses known to possess a genome consisting of circular double-stranded dna. we have characterized the viral transcript and determined the complete sequence of the genome of commelina mellow mottle virus (coymv), a member of this group. analysis of the viral transcript indicates that the virus encodes a single terminally-redundant genome-length plus 120 nucleotide transcript. a fraction of the transcripts is polyadenylated, alt ...19901699203
activation of a truncated pr-1 promoter by endogenous enhancers in transgenic plants.pr-1 genes are induced by various environmental stimuli such as pathogen attack or exposure of the plants to certain chemicals. to examine the regulation of these genes, the 5' flanking regions of the pr-la gene and of two pr-1 pseudogenes were joined by a transcriptional fusion to the escherichia coli beta-glucuronidase (gus) gene. these constructs were stably integrated into the tobacco genome and independent primary transformants were monitored for the expression of the reporter gene. unexpec ...19921731979
new patterns of gene activity in plants detected using an agrobacterium vector.a vector has been designed that contains a truncated camv (cauliflower mosaic virus) 35s promoter fused to a receptor gene encoding beta-glucuronidase (gus), placed adjacent to the left border sequence of an agrobacterium vector. in potato plants transformed with this vector, different patterns of transcription were detected at high frequency using in situ assays for gus activity. previous studies in drosophila using analogous vectors have shown that the new patterns of transcription in many cas ...19911893100
cdnas of beet necrotic yellow vein virus rnas 3 and 4 are rendered biologically active in a plasmid containing the cauliflower mosaic virus 35s promoter.cdnas of beet necrotic yellow vein virus rnas 3 and 4 could be rendered biologically active when they were placed under the control of the cauliflower mosaic virus 35s promoter and polyadenylation signal. although the 35s in vivo transcripts should have contained up to forty 5' and several hundred 3' nonviral nucleotides, the progeny viral rnas had the same sizes as in naturally infected sugarbeets. the progeny rnas did not hybridize with the nonviral sequences indicating that they were apparent ...19911926790
translation of a polycistronic mrna in the presence of the cauliflower mosaic virus transactivator protein.polycistronic mrnas containing an upstream beta-glucuronidase (gus) and a downstream chloramphenicol acetyltransferase (cat) reporter open reading frame (orf) were expressed in transfected plant protoplasts. cat expression could be strongly induced by coexpression of the cauliflower mosaic virus encoded translation transactivator. transactivation was abolished when an upstream orf overlapped the cat orf for a long distance. no specific sequence elements were required for transactivation but the ...19911935908
effect of deletions in the cauliflower mosaic virus polyadenylation sequence on the choice of the polyadenylation sites in tobacco protoplasts.deletions were made in the cauliflower mosaic virus polyadenylation sequence which was cloned downstream of the beta-glucuronidase gene (gus). the populations of mrnas generated in tobacco mesophyll protoplasts by transient expression with the various constructs were analysed using a polymerase chain reaction procedure. when no deletion was present in the sequence, the mrna appeared to be polyadenylated at two major polyadenylation sites. a deletion upstream from the aataaa sequence made the pop ...19911709718
the cauliflower mosaic virus reverse transcriptase is not produced by the mechanism of ribosomal frameshifting in saccharomyces cerevisiae.the capsid protein and the reverse transcriptase of cauliflower mosaic virus (camv) are encoded by two genes (orf iv and orf v) that lie in different translation reading frames. a comparison can be drawn between the synthesis of both camv proteins and the fusion protein in a yeast retrotransposon, ty, resulting from a +1 frameshifting event which fuses two out-of-phase orfs encoding the structural protein and the reverse transcriptase of ty. for this reason, we constructed a yeast expression vec ...19911703375
subcellular location and expression level of a chimeric protein consisting of the maize waxy transit peptide and the beta-glucuronidase of escherichia coli in transgenic potato plants.the transit peptide of the maize waxy protein (a nuclear-encoded amyloplast protein of the maize endosperm) was studied with respect to its role in subcellular protein targeting in transgenic potato plants. tp30, a chimeric precursor protein consisting of the waxy transit peptide and an additional 34 amino acids of the mature waxy protein fused to the beta-glucuronidase of escherichia coli, was expressed in potato plants under the control of the 35s promoter of cauliflower mosaic virus. this fus ...19912005871
in vitro dna methylation inhibits gene expression in transgenic tobacco.a hemimethylated chimeric gene, containing the cauliflower mosaic virus 35s promoter, the beta-glucuronidase coding region and the polyadenylation signal of nopaline synthase, was introduced into tobacco protoplasts by polyethylene glycol mediated transfection. hemimethylation led to complete inhibition of transient gene expression. in regenerated transgenic plants the integrated gene was constitutively hypermethylated at the sequences cpg and cpnpg and this was correlated with an inactivation o ...19901702383
functional analysis of the pathogenesis-related 1a protein gene minimal promoter region. comparison of reporter gene expression in transient and in stable transfections.pathogenesis-related (pr) proteins are a heterogeneous group of host encoded, low-molecular-mass proteins that are induced in plants by various external stimuli, such as pathogen attack or exposure of the plants to certain chemicals. to examine the regulation of these genes, the 5'-flanking region of the tobacco pr-1a gene [pfitzner u.m., pfitzner, a.j.p. & goodman, h.m. (1988) mol. gen. genet. 211, 290-295] was joined by a transcriptional fusion to the escherichia coli beta-glucuronidase (gus) ...19912007405
expression of cowpea mosaic virus coat protein precursor in transgenic tobacco plants.tobacco, nicotiana tabacum l., supports cowpea mosaic virus (cpmv) replication and cell-to-cell movement, and thus may serve as a model system to study coat protein-mediated protection against cpmv. a chimeric gene consisting of the cauliflower mosaic virus 35s promoter, cpmv 60k coat proteins-precursor (cp-p) coding region, and the nopaline synthase polyadenylation signal was transferred to tobacco cv. burley 21 via the agrobacterium tumefaciens binary vector system. gene integration and expres ...19921730936
targeting of glutamine synthetase to the mitochondria of transgenic tobacco.two transgenic tobacco lines were genetically engineered to contain chimaeric genes encoding the glutamine synthetase (gs) gamma polypeptide of phaseolus vulgaris (french bean), expressed from the cauliflower mosaic virus 35s promoter. one (mit-1) contained two copies of a construct including the first 60 amino acids of the nicotiana plumbaginifolia beta-f1 atpase to target the gs polypeptide to the mitochondrion. the other (cyt-4) contained a single copy of a cytosolic gs construct. leaves of i ...19901983302
barley aleurone layer cell protoplasts as a transient expression system.protoplasts were prepared from barley aleurone layers using 'onozuka' cellulase digestion and purification through a percoll gradient. protoplasts prepared by this procedure had a viability ranging from 60% to 80% during the first two days of culture. they were responsive to gibberellic acid (ga) as measured by the stimulation of alpha-amylase synthesis. the ga stimulation was counteracted by abscisic acid (aba). in the presence of polyethylene glycol (peg), the protoplasts took up exogenously a ...19911832576
upstream sequences other than aauaaa are required for efficient messenger rna 3'-end formation in plants.we have characterized the upstream nucleotide sequences involved in mrna 3'-end formation in the 3' regions of the cauliflower mosaic virus (camv) 19s/35s transcription unit and a pea gene encoding ribulose-1,5-bisphosphate carboxylase small subunit (rbcs). sequences between 57 bases and 181 bases upstream from the camv polyadenylation site were required for efficient polyadenylation at this site. in addition, an aauaaa sequence located 13 bases to 18 bases upstream from this site was also impor ...19901983794
regulation of a modified camv 35s promoter by the tn10-encoded tet repressor in transgenic tobacco.we have investigated the use of the tn10-encoded tet repressor-operator system to regulate the expression of a suitably engineered cauliflower mosaic virus (camv) 35s promoter in transgenic tobacco plants. first, a transgenic plant was generated which constitutively synthesizes 600,000 tet repressor monomers per cell. in a second transformation step, the beta-glucuronidase (gus) gene under the control of a modified camv 35s promoter, containing two tet operators, was stably integrated into the p ...19912062303
high-level expression of a tobacco chitinase gene in nicotiana sylvestris. susceptibility of transgenic plants to cercospora nicotianae infection.endochitinases (e.c.3.2.14, chitinase) are believed to be important in the biochemical defense of plants against chitin-containing fungal pathogens. we introduced a gene for class i (basic) tobacco chitinase regulated by cauliflower mosaic virus 35s-rna expression signals into nicotiana sylvestris. the gene was expressed to give mature, enzymatically active chitinase targeted to the intracellular compartment of leaves. most transformants accumulated extremely high levels of chitinase-up to 120-f ...19911888892
tissue-specific expression of the tmv coat protein in transgenic tobacco plants affects the level of coat protein-mediated virus protection.transgenic tobacco plants were produced that express a chimeric gene encoding the coat protein (cp) of tobacco mosaic virus (tmv) under the control of the promoter from a ribulose bisphosphate carboxylase small subunit (rbcs) gene. plant lines expressing comparable levels of cp from the rbcs and cauliflower mosaic virus 35s promoters were compared for resistance to tmv. in whole plant assays the 35s:cp constructs gave higher resistance than the rbcs:cp constructs. on the other hand, leaf mesophy ...19902238465
characteristics of a strong promoter from figwort mosaic virus: comparison with the analogous 35s promoter from cauliflower mosaic virus and the regulated mannopine synthase promoter.a segment of dna from the genome of figwort mosaic virus (fmv) strain m3 possesses promoter activity when tested in electroporated protoplasts from, and transgenic plants of, nicotiana tabacum cv. xanthi nc. the 1.1 kb dna segment, designated the '34s' promoter, is derived from a position on the fmv genome comparable to the position on the cauliflower mosaic virus (camv) genome containing the 35s promoter. the 34s and 35s promoters show approximately 63% nucleotide homology in the tata, ccact, a ...19902102823
quantitation of chloramphenicol acetyl transferase in transgenic tobacco plants by elisa and correlation with gene copy number.a monoclonal antibody to chloramphenicol acetyl transferase (cat) was used in an indirect competitive enzyme immunoassay (elisa) for the quantitation of cat in leaf extracts of eighteen transgenic tobacco plants containing the cat gene fused to the cauliflower mosaic virus 35s promoter. the elisa could be used to quantify cat when present in extracts at 20 ng/ml. enzymatic activity and electrophoretic mobility of cat in these extracts was not different from cat from escherichia coli. concentrati ...19902102836
replication of a geminivirus derived shuttle vector in maize endosperm cells.a maize (zea mays l.) endosperm cell culture has been shown to efficiently replicate dna sequences derived from wheat dwarf virus (wdv), a monopartite monocot geminivirus. to analyze sequences necessary for viral replication and to verify their application for a plant gene expression vector, we have developed a 3.7 kilobase pairs escherichia coli--plant cell shuttle vector, pwi-11. the p15a origin of replication, functional in e. coli, was introduced into the viral sequences. we have replaced th ...19911849629
histochemical analysis of camv 35s promoter-beta-glucuronidase gene expression in transgenic rice plants.the cauliflower mosaic virus promoter is commonly used to drive transcription of chimeric genes in transgenic plants, including the cereals. to determine the tissue and cell types of cereal plants that the promoter functions in, transgenic rice plants containing a camv 35s promoter/gus chimeric gene were analyzed for gus activity. insertion of a 35s/gus chimeric gene at low copy number into chromosomal dna of plants regenerated from electroporated protoplasts was confirmed by gel blot hybridizat ...19902102372
molecular analysis of two pr-1 pseudogenes from tobacco.two independent pr-1 lambda genomic clones (w38/1 and w38/3) were isolated and characterized from a tobacco (nicotiana tabacum cv. wisconsin 38) library. neither clone is identical to the previously described pr-1 cdna clones, and both clones carry mutations within the highly conserved pr-1 protein coding region. for example, clone w38/1 has a gaa glu codon instead of the translation stop codon thus harbouring an open reading frame extended by 16 additional amino acids. furthermore, both clones ...19911888891
analysis of tomato polygalacturonase expression in transgenic tobacco.tomato polygalacturonase is a cell wall enzyme secreted in large amounts during tomato fruit ripening. polygalacturonase is synthesized as a glycoprotein precursor that undergoes numerous cotranslational and post-translational processing steps during its maturation, yielding three isozymes in tomato fruit, pg1, pg2a, and pg2b. to investigate the physiological roles of the three isozymes and the functional significance of the polygalacturonase processing domains in its intracellular transport and ...19902152163
the sv40 small t intron is accurately and efficiently spliced in tobacco cells.we have introduced the sv40 small t intron into tobacco cells as part of a cauliflower mosaic virus 35s promoter-chloramphenicol acetyltransferase-sv40 transcription unit. we find that the small t intron is efficiently and accurately spliced in transgenic tobacco cells that carry this transcription unit. our results indicate that there is substantial conservation of rna processing signals between plants and animals, more than has been previously assumed. they also suggest that pre-mrna processin ...19911654158
transfer of methomyl and hmt-toxin sensitivity from t-cytoplasm maize to tobacco.the mitochondrial gene, t-urf13, which is unique to the t-cytoplasm of maize, has been expressed in tobacco plants using the cauliflower mosaic virus 35s promoter. tobacco plants expressing t-urf13 exhibit a variety of responses to methomyl. leaf discs and petiole sections bleach when exposed to methomyl or hmt-toxin; this effect increases with the age of the tissue. the bleaching effect is not however observed when light is excluded. plants homozygous for t-urf13 exhibit extreme sensitivity whe ...19911944229
kinetics of tomato golden mosaic virus dna replication and coat protein promoter activity in nicotiana tabacum protoplasts.we have analyzed the replication kinetics of the dna a and dna b genome components of the geminivirus tomato golden mosaic virus (tgmv) in protoplasts derived from nicotiana tabacum suspension culture. in addition, the kinetics of tgmv coat protein promoter activity, as measured by expression of a beta-glucuronidase (gus) reporter, have been examined. in our protoplast system, double-stranded dna forms of both viral genome components appeared by 18 hr post-transfection, while single-stranded dna ...19921736521
custom polymerase-chain-reaction engineering of a plant expression vector.polymerase-chain-reaction (pcr) amplification combined with custom-synthesized oligodeoxyribonucleotide (oligo) primers can be used to make complex genetic engineering steps (e.g., translational fusions) easy. much of the complexity of the engineering steps can be incorporated into the custom oligo primers. using this technique, a plant constitutive expression vector, puc18cpexp, was constructed. this vector is based on the cauliflower mosaic virus 35s gene-regulatory elements and the cucumber m ...19912055473
role of propeptide glycan in post-translational processing and transport of barley lectin to vacuoles in transgenic tobacco.mature barley lectin is a dimeric protein composed of two identical 18-kilodalton polypeptides. the subunits of barley lectin are initially synthesized as glycosylated proproteins, which are post-translationally processed to the mature protein preceding or concomitant with deposition of barley lectin in vacuoles. to investigate the functional role of the glycan in processing and intracellular transport of barley lectin to vacuoles, the sole n-linked glycosylation site residing within the cooh-te ...19902152118
analysis of regulatory elements involved in the induction of two tobacco genes by salicylate treatment and virus infection.tobacco genes encoding the pr-1a protein and a glycine-rich protein are expressed after treatment of plants with salicylate or infection with tobacco mosaic virus. upstream sequences of these genes were fused to reporter genes, and these constructs were used to transform tobacco. upstream sequences of the pr-1a gene of 689 base pairs or longer were sufficient for induction of the reporter gene in tobacco mosaic virus-inoculated leaves, systemically induced leaves from infected plants, and leaves ...19902152122
a metal-dependent dna-binding protein interacts with a constitutive element of a light-responsive promoter.we have used dnase i footprinting to characterize nuclear factors that bind to the light-responsive promoter of pea rbcs-3a, one member of the gene family encoding the small subunit of ribulose-1,5-bisphosphate carboxylase. a sequence-specific binding activity, designated 3af1, binds to an at-rich sequence present at the -45 region of the rbcs-3a promoter. a tetramer of the 3af1 binding site, designated as box vi, can form multiple complexes with tobacco leaf and root nuclear extracts. mutations ...19902152132
disease symptoms in transgenic tobacco induced by integrated gene vi of cauliflower mosaic virus.a chimeric vector (pkr 612b1) containing the neomycin phosphotransferase (aph) gene from the tn5 transposon under the control of the gene vi promoter of cauliflower mosaic virus (camv) and the cloned gene vi region (sali-bsteii) of the same virus were used to cotransform tobacco protoplasts. using the polyethylene glycol transformation procedure, a large number of protoplasts were transformed and proved to be resistant to kanamycin (km). whole km-resistant plants were regenerated and shown to co ...19902161156
analysis of acidic and basic chitinases from tobacco and petunia and their constitutive expression in transgenic tobacco.cdna clones of messenger rnas for acidic and basic chitinases were isolated from libraries of tobacco mosaic virus-infected samsun nn tobacco and petunia. the tobacco cdna clones for acidic chitinase fell into two different groups, whereas all petunia cdna clones had the same sequence. also, tobacco genomic clones were isolated and one was characterized. this genomic clone, corresponding to one of the cdna clones, showed that this acidic chitinase gene contains two introns. the amino acid sequen ...19902131096
analysis of unstable rna transcripts of insecticidal crystal protein genes of bacillus thuringiensis in transgenic plants and electroporated protoplasts.we have examined expression of several insecticidal crystal protein (icp) genes of bacillus thuringiensis in transgenic tobacco plants and electroporated carrot protoplasts. we determined that low levels of lepidopteran toxin cryia(b) icp gene expression in plants and electroporated carrot cells is due to rna instability. we used a series of 3' deleted by cryia(b) constructs directed by the cauliflower mosaic virus 35s promoter to demonstrate that this instability is minimally contained in the f ...19911863758
vacuolar targeting and posttranslational processing of the precursor to the sweet potato tuberous root storage protein in heterologous plant cells.sporamin, the tuberous root storage protein of the sweet potato, which is localized in vacuoles, is synthesized as a prepro-precursor with an n-terminal sequence of amino acids that includes a signal peptide and an additional pro-segment of 16 amino acids. a full-length cdna for sporamin was placed downstream of the 35 s promoter of cauliflower mosaic virus and introduced into tobacco and sunflower genomes by ti plasmid-mediated transformation. a polypeptide of nearly the same size as mature spo ...19902246259
the cauliflower mosaic virus open reading frame vii product can be expressed in saccharomyces cerevisiae but is not detected in infected plants.antiserum was prepared against a synthetic peptide corresponding to the n-terminal 20 amino acids of the protein encoded by cauliflower mosaic virus (camv) open reading frame vii (orf vii). this antiserum was used to detect the expression of camv orf vii either in saccharomyces cerevisiae transformed by an expression vector containing camv orf vii or in camv-infected plants. only in s. cerevisiae has a 14-kilodalton protein been detected.19902186173
propeptide of a precursor to a plant vacuolar protein required for vacuolar targeting.sporamin is a protein without glycans that accumulates in large quantities in the vacuoles of the tuberous root of the sweet potato. it is synthesized as a prepro precursor with an n-terminal extension composed of a 21-amino-acid signal peptide and a 16-amino-acid propeptide. a total of 48 base pairs, corresponding to the nucleotide sequence that encodes the propeptide, was deleted from a cdna clone for sporamin. this delta pro mutant sequence, as well as the sequence of the wild-type sporamin c ...19911992474
expression of cauliflower mosaic virus gene i using a baculovirus vector based upon the p10 gene and a novel selection method.a new baculovirus expression vector based upon the p10 gene of autographa californica nuclear polyhedrosis virus (acnpv) and a novel system for the screening of p10 recombinants have been developed. the insertion of a cassette containing the lacz gene under the control of a heat-shock promoter of drosophila melanogaster downstream from the cloning site in p10 transfer vectors allows the convenient identification of putative recombinants by virtue of their expression of beta-galactosidase. using ...19902219726
expression of cauliflower mosaic virus gene i in insect cells using a novel polyhedrin-based baculovirus expression vector.an improved polyhedrin-based baculovirus expression vector was constructed to expedite distinguishing infections by putative baculovirus recombinants from infections by wild-type (wt) baculovirus. the vector utilizes the escherichia coli beta-galactosidase gene (lacz) as a genetic marker for positive recombination between wt autographa californica nuclear polyhedrosis virus and the baculovirus transfer vector. the marker gene/expression cassette was constructed so that lacz and the deleted polyh ...19902230725
recognition efficiency of dicotyledoneae-specific promoter and rna processing signals in rice.heterologous gene expression experiments have shown that genes of monocotyledoneae are often not transcribed in dicotyledoneae, or produce pre-mrna that is inefficiently or aberrantly processed. it is however not known how correctly and efficiently dicotyledon-specific gene expression signals are recognized in cells of monocotyledoneae. here we address this question using tobacco (nicotiana tabacum) and rice (oryza sativa) protoplasts transformed with the same hybrid gene constructs. constructs ...19902177137
design and cloning of a synthetic gene for the flounder antifreeze protein and its expression in plant cells.a synthetic gene coding for the winter flounder antifreeze protein (afp) has been constructed. a new strategy for the synthesis has been employed such that one strand of the duplex was chemically synthesized and the other was produced enzymatically by chain extension. the chemically synthesized blocks were constructed so that the second strand was self-priming. the resulting dna fragment was incorporated into the vector, pgcs1, which contained a translational fusion of the sequence encoding afp ...19902210378
effect of cpg methylation on gene expression in transfected plant protoplasts.activity of the cat gene driven by the cauliflower mosaic virus 35s promoter has been assayed by transfecting petunia protoplasts with the puc8camvcat plasmid. in vitro methylation of this plasmid with m.hpaii (methylates c in ccgg sites) and m.hhai (methylates gcgc sites) did not affect bacterial chloramphenicol acetyltransferase (cat) activity. it should be noted, however, that no hpaii or hhai sites are present in the promoter sequence. in contrast, in vitro methylation of the plasmid with th ...19902258051
enhancement of foreign gene expression by a dicot intron in rice but not in tobacco is correlated with an increased level of mrna and an efficient splicing of the intron.the first intron of castor bean catalase gene, cat-1 was placed in the n-terminal region of the coding sequence of the beta-glucuronidase gene (gusa) and the intron-containing gusa was used with the cauliflower mosaic virus (camv) 35s promoter. using this plasmid, pig221, the effect of the intron on expression of beta-glucuronidase (gus) activity was examined in transgenic rice calli and plants (a monocotyledon), and transgenic tobacco plants (a dicotyledon). the intron-containing plasmid increa ...19902263444
infectivity of plasmids containing brome mosaic virus cdna linked to the cauliflower mosaic virus 35s rna promoter.full-length biologically active cdnas of brome mosaic virus genomic rnas 1, 2 and 3 were constructed by joining cdna fragments. the cdnas were constructed so that, at the 5' ends, unique snabi sites were present at the site of initiation of transcription. the cdnas were inserted between a modified cauliflower mosaic virus (camv) 35s rna promoter and terminator regions derived from camv dna, and cloned into puc18. when a mixture of the plasmid dnas was inoculated onto chenopodium hybridum leaves, ...19911993867
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