| the restriction endonucleases in bacillus amyloliquefaciens n strain. substrate specificities. | two species of restriction endonuclease were isolated by gel filtration and deae-cellulose chromatography from a cell-free extract of bacillus amyloliquefaciens (b. subtilits) n strain; a lower molecular weight endonuclease (endonuclease r.bamni) and a higher molecular-weight one (endonuclease r.bamnx). both of them required only mg2+ for their activities. endonuclease r.bamnx introduced a larger number of site-specific scissions in excherchia coli phage lambda dna that endonuclease r.bamni did. ... | 1976 | 182257 |
| purification and properties of an endonuclease from nuclei of uninfected and polyoma-infected 3t3 cells. | an endonuclease activity has been purified approximately 800-fold from nuclei of 3t3 cells infected with polyoma virus. the purfied enzyme catalyzes an endonucleoytic cleavage of single- and double-stranded dna and single-stranded rna. evidence that the activity towards these substrates resides in the same protein molecule is provided by the finding that they co-sediment in sucrose gradients and have identical rates of heat inactivation. studies on the dnase activity shows that the rate of hydro ... | 1976 | 187596 |
| restriction enzyme cleavage mapping of t7 virus early region. | dna molecules of seven t7 mutants with overlapping deletions in the early region were cleaved by restriction enzymes hindii, hpai and ii, and haeiii. the differences in the cleavage patterns after electrophoresis have been used to generate a cleavage map of the restriction sites of this enzyme. it covers the first 9% of the t7 dna molecule. cleavage points for hindii are at 0.60, 1.33, 1.59, 1.76, 5.26, 6.27, 7.4 and 8.38%; for hpai and ii at 1.36, 1.62, 4.46, 6.29, 6.62, 7.56, and 8.76%, for ha ... | 1978 | 355837 |
| is the dna of virus t7 methylated? | dna modification of t7 wild type and of t7 m-mutants was studied by determining the percentage of 5-methylcytosine (5mc)/cytosine (c) and n6-aminopurine (6ma)/adenine (a) and by evaluating the plating efficiencies of restriction-sensitive t7 m-mutants on modifying and non-modifying host strains. only 0.03% adenine and 0.02% cytosine were methylated in the dna of t7 wild type as well as in t7 m-mutants, which was independent of the host dna methylation (30- to 50-fold higher). the restriction of ... | 1979 | 393804 |
| a virus-specified mechanism for the prevention of multiple infection--t7- and t3-mutual and superinfection exclusion. | co- and superinfection of cells with t3/t7 result in exclusion (mutual or superinfection exclusion). the exclusion mechanism is also directed against homologous (or identical) virus. exclusion is established after the adsorption but before the genome becomes available for gene expression or replication, that is only one virus per cell develops. the exclusion is triggered by a constituant of the viral particle. an early t7 gene (m gene) (schweiger et al., 1975) is essential for the formation of e ... | 1976 | 799243 |
| helper function of t7 protein kinase in virus propagation. | | 1975 | 1094300 |
| permeability lesions in male escherichia coli infected with bacteriophage t7. | the abortive development of bacteriophage t7 in e. coli cells carrying f factors has previously been attributed to a lack of virus-directed modification of ribosomes in such cells. we find it unnecessary to postulate such translational control to explain the failure of t7 development. instead, there is a general cessation of macromolecular syntheses around 8 min after t7 infection of f' cells. this cessation is correlated with a sudden outflow of the entire acid-soluble pool of phosphorus-contai ... | 1975 | 1094459 |
| molecular weight of dna from actinophage msp2. | actinophage msp2 is infectious for streptomyces venezuelae s13. based upon electron microscopy of coliphage t4 mixed with msp2, msp2 had a head about 48 +/- 2 nm wide and 87 +/- 5 nm long. dna from polyoma virus and from coliphages t4 and t7 served as reference markers in estimating the molecular weight of msp2 dna from sedimentation in sucrose gradients. denatured msp2 dna was estimated to be about 17 x 10(6) and double-stranded msp2 dna was about (36 +/- 1.6) x 10(6) in molecular weight. | 1975 | 1159900 |
| beta-l-thymidine 5'-triphosphate analogs as dna polymerase substrates. | beta-l-3'-deoxythymidine 5'-triphosphate (l-ddttp) and beta-l-3'-deoxy-2',3'-didehydrothymidine 5'-triphosphate (l-d4ttp) were substrates for human immunodeficiency virus reverse transcriptase, escherichia coli dna polymerase i (klenow), and sequenase (modified t7 dna polymerase). the beta-d- and beta-l-enantiomers of 5-methyluridine 5'-triphosphate (rttp) were inhibitors but not substrates of reverse transcriptase. the steady-state km values for l-ddttp and l-d4ttp, with all three enzymes, were ... | 1992 | 1281153 |
| duck hepatitis b virus polymerase produced by in vitro transcription and translation possesses dna polymerase and reverse transcriptase activities. | activities of the hepadnavirus polymerases are known to include those of dna polymerase, reverse transcriptase and rnase h. to date, it has been difficult or impossible to clone and express the product as an active enzyme. in this study, full length capped rna encoding duck hepatitis b virus (dhbv) polymerase was produced by in vitro transcription from a t7 promoter. the rna was translated in a rabbit reticulocyte lysate system and produced an 35s-methionine labelled 79 kd band on sds-polyacryla ... | 1992 | 1281990 |
| the 9-kda hydrophobic protein encoded at the 3' end of the porcine transmissible gastroenteritis coronavirus genome is membrane-associated. | the open reading frame potentially encoding a 78 amino acid, 9101 da hydrophobic protein (hp) and, mapping at the 3' end of the porcine transmissible gastroenteritis coronavirus (tgev) genome, was shown to be expressed during virus replication. the cloned hp gene was placed in a plasmid under control of the t7 rna polymerase promoter and in vitro translation of transcripts generated in vitro yielded a 9.1-kda protein that was immunoprecipitable with porcine hyperimmune anti-tgev serum. antiserum ... | 1992 | 1310191 |
| internal ribosome entry site within hepatitis c virus rna. | the mechanism of initiation of translation on hepatitis c virus (hcv) rna was investigated in vitro. hcv rna was transcribed from the cdna that corresponded to nucleotide positions 9 to 1772 of the genome by using phage t7 rna polymerase. both capped and uncapped rnas thus transcribed were active as mrnas in a cell-free protein synthesis system with lysates prepared from hela s3 cells or rabbit reticulocytes, and the translation products were detected by anti-gp35 antibodies. the data indicate t ... | 1992 | 1310759 |
| expression of cdna encoding the sendai virus hemagglutinin-neuraminidase gene: characterization of wild-type and mutant gene products. | cloned cdna encoding the sendai virus (sv) hemagglutinin-neuraminidase (hn) envelope glycoprotein was expressed in cultured cells in two ways: (i) infection with hn-expressing recombinant vaccinia virus, or (ii) transfection with a plasmid with t7 promoter and termination sequences flanking the hn gene, with intracellular t7 rna polymerase supplied by coinfection with recombinant vaccinia virus that expresses the enzyme. the hn expressed was indistinguishable from the authentic sv protein in ant ... | 1992 | 1312281 |
| globic: a very fast microcomputer program for fingerprinting, characterization and comparison of long nucleotide sequences. | this paper describes the program globic, which compares, characterizes and fingerprints even 0.1 mbase sequences in a few minutes with the aid of an ibm-at microcomputer. instead of the nucleotide sequences themselves, globic compares the local nucleotide or short oligonucleotide compositions. globic presents two-dimensional maps of contour lines depicting the similarity of two different sequences, a sequence compared to itself, to its complementary sequence or to a random sequence. a vocabulary ... | 1992 | 1314685 |
| transient expression and mutational analysis of the rotavirus intracellular receptor: the c-terminal methionine residue is essential for ligand binding. | maturation of rotavirus involves an intracellular membrane budding event in which the single-shelled icosahedral particle interacts with a virus-encoded receptor glycoprotein, ns28, that is located in the rough endoplasmic reticulum membrane. the receptor is a tetramer and is oriented with the c-terminal 131 amino acids on the cytoplasmic side of the membrane (a.r. bellamy and g.w. both, adv. virus res. 38:1-48, 1990). we have used the t7-vaccinia virus transient expression system to deliver mut ... | 1992 | 1316468 |
| localization of an rna-binding domain in the nucleocapsid protein of the coronavirus mouse hepatitis virus. | the interaction between the nucleocapsid (n) protein of mouse hepatitis virus (mhv) and rna was studied in an effort to define portions of the n molecule that participate in binding to rna. n mrnas transcribed from sp6 and t7 vectors were translated in a rabbit reticulocyte lysate. analysis of synthesized n protein in a nondenaturing gel system showed that it bound in vitro to an endogenous rna in the reticulocyte lysate but not to its own mrna. a set of deletion mutants was constructed in order ... | 1992 | 1322650 |
| mutational analysis of the proposed fg loop of poliovirus proteinase 3c identifies amino acids that are necessary for 3cd cleavage and might be determinants of a function distinct from proteolytic activity. | mutations were introduced into a cdna clone of poliovirus resulting in single-amino-acid substitutions within the region of the proposed fg loop of proteinase 3c. rnas were made by in vitro transcription with t7 rna polymerase and used to transfect hela cells. virus viability was assessed as indicated by cell lysis. in parallel, rnas were translated in vitro by using a hela cell lysate, and the patterns of the processed poly-proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel ele ... | 1992 | 1326654 |
| molecular cloning of natural paramyxovirus copy-back defective interfering rnas and their expression from dna. | using the unique sequence organization of copy-back defective interfering (di) rnas of paramyxoviruses, sendai virus (sv), and measles virus copy-back di rnas were pcr amplified and cloned, without having to separate them from their helper nondefective genomes. the cloning was designed so that t7 polymerase transcription of the plasmids would generate di rnas with the exact 5' and 3' ends. the sv di clone, transcribed from the plasmid in bhk cells using t7 polymerase produced by a vaccinia virus ... | 1992 | 1329337 |
| cloning and expression of a cdna encoding the epstein-barr virus thymidine kinase gene. | a clone of the epstein-barr virus (ebv) thymidine kinase (tk) gene was derived from a cdna library of p3hr1 cells. the gene product was expressed as a fusion protein in a procaryotic system by using t7 rna polymerase. the recombinant tk showed a molecular mass of 67 kda and was biologically active. antiserum raised in mice immunized with partially purified tk recognized an antigen present in ebv-superinfected raji cells using an indirect immunofluorescence assay. | 1992 | 1331157 |
| multiple glycoproteins synthesized by the smallest rna segment (s10) of bluetongue virus. | the genome of bluetongue virus, an orbivirus, consists of 10 double-stranded rnas, each encoding at least one polypeptide. the smallest rna segment (s10) encodes two minor nonstructural proteins, ns3 and ns3a, the structures and functions of which are not understood. we have expressed these two proteins in mammalian cells by using the t7 cytoplasmic transient expression system. using a deletion mutant (lacking the first aug initiation codon), we have demonstrated that the second initiation codon ... | 1992 | 1331513 |
| fidelity of human immunodeficiency virus type i reverse transcriptase in copying natural rna. | the in vitro fidelity of reverse transcriptase from human immunodeficiency virus type i (hiv-1 rt) upon copying an rna template was measured using the phi xam 16 reversion assay. a phi x174 sequence harboring the amber 16 codon was cloned into a transcription vector. rna obtained from transcription by bacteriophage t7 rna polymerase was used as a template for rna-directed dna synthesis by hiv-1 rt. an imbalance of dntp concentrations during the reverse transcription step served to distinguish be ... | 1992 | 1371812 |
| mechanism of interferon action: cdna structure, expression, and regulation of the interferon-induced, rna-dependent p1/eif-2 alpha protein kinase from human cells. | a molecular cdna clone (p1 kin) was isolated that encodes the human rna-dependent p1/eif-2 alpha protein kinase. the complete cdna sequence of the p1 kin cdna was determined; the longest open reading frame (orf) encoded a 551 amino acid protein with a deduced molecular weight of 62055 da. transcripts prepared from the p1 kin cdna by transcription in vitro with t7 rna polymerase programmed the cell-free synthesis of a protein indistinguishable by immunoprecipitation and immunoblot gel analyses fr ... | 1992 | 1373553 |
| 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 ... | 1992 | 1374981 |
| complete replication of a satellite rna in vitro by a purified rna-dependent rna polymerase. | the 334 nucleotide r satellite rna was used as a template for purified rna-dependent rna polymerase (rdrp) from cucumber mosaic virus-infected tobacco plants. the products of the reaction were dsrna and positive-strand rna of the same size as the r satellite rna. similar products were obtained when t7 rna polymerase positive-strand transcripts of a cdna clone of the satellite rna, designed to have the same 5' and 3' ends as the satellite rna, were used as templates. the formation of the positive ... | 1992 | 1376771 |
| unequal human immunodeficiency virus type 1 reverse transcriptase error rates with rna and dna templates. | sequence variation in the type 1 human immunodeficiency virus (hiv-1) results, in part, from inaccurate replication by reverse transcriptase. although this enzyme is error-prone during synthesis in vitro with dna templates, the fidelity of rna-dependent dna synthesis relevant to minus-strand replication in the virus life cycle has not been examined extensively. in the present study, we have developed a system to determine the fidelity of transcription and reverse transcription and have used it t ... | 1992 | 1379727 |
| interaction of initiation factors with the cap structure of chimaeric mrna containing the 5'-untranslated regions of semliki forest virus rna is related to translational efficiency. | chimaeric chloramphenicol acetyltransferase (cat) mrna, containing the leader sequences of genomic 42s rna and subgenomic 26s rna of semliki forest virus (sfv) were synthesized by in-vitro transcription. these transcripts were translated with different efficiencies, as the authentic mrna in sfv-infected cells. therefore, they can be used as model mrna species to study the mechanism underlying sfv-directed shut off of host protein synthesis. the interaction of translation initiation factors with ... | 1992 | 1396664 |
| expression of functional parvoviral ns1 from recombinant vaccinia virus: effects of mutations in the nucleotide-binding motif. | the gene encoding the major replicative protein, ns1, of minute virus of mice (mvm) was transferred into a recombinant vaccinia virus vector in place of the vaccinia thymidine kinase gene. the ns1 gene was placed under control of a bacteriophage t7 promoter and expressed in cells coinfected with another recombinant vaccinia virus, vtf7-3, which encodes the t7 rna polymerase. expression of ns1 was further enhanced by the presence of a 5' untranslated region, derived from encephalomyocarditis viru ... | 1992 | 1413512 |
| cytoplasmic delivery of ribozymes leads to efficient reduction in alpha-lactalbumin mrna levels in c127i mouse cells. | ribozymes targeted to five sites along the alpha-lactalbumin (alpha-lac) mrna were delivered to the cytoplasm of mouse c127i mammary cells using the t7-vaccinia virus delivery system and the amount of alpha-lac mrna was monitored 24-48 h post-transfection. three target sites were selected in the alpha-lac coding region (nucleotides 15, 145 and 361) and two were located in the 3' non-coding region (nucleotides 442 and 694). acting in trans and at a target:ribozyme ratio of 1:1000, ribozymes targe ... | 1992 | 1425576 |
| a vector, psht, for the expression and secretion of protein domains in mammalian cells. | a phagemid (psht) containing the puc and m13 ori sequences was constructed to facilitate the expression of partial cdnas or of sequences encoding mammalian membrane- and secretory-protein domains. it provides a start codon and signal sequence flanked upstream by the simian virus 40 and bacteriophage t7 promoters and downstream by cloning sites, stop codons in all three frames, splicing and polyadenylation signals. | 1992 | 1427094 |
| transmembrane signaling by the human insulin receptor kinase. relationship between intramolecular beta subunit trans- and cis-autophosphorylation and substrate kinase activation. | to examine the role of intramolecular beta subunit trans- and cis-autophosphorylation in signal transduction, the vaccinia virus/bacteriophage t7 expression system was used to generate insulin holoreceptors composed of a kinase-defective half-receptor precursor (alpha beta a/k or alpha beta a/k.delta ct) and a kinase-active half-receptor precursor (alpha beta delta ct or alpha beta wt). in the alpha beta a/k-alpha beta delta ct hybrid insulin receptor, insulin stimulated a 20-fold increase in in ... | 1992 | 1326556 |
| importance of p12 protein in mason-pfizer monkey virus assembly and infectivity. | mason-pfizer monkey virus (m-pmv) represents the prototype type d retrovirus, characterized by the assembly of intracytoplasmic a-type particles within the infected-cell cytoplasm. these immature particles migrate to the plasma membrane, where they are released by budding. the gag gene of m-pmv encodes a novel protein, p12, just 5' of the major capsid protein (ca) p27 on the polyprotein precursor. the function of p12 is not known, but an equivalent protein is found in mouse mammary tumor virus a ... | 1992 | 1433505 |
| molecular characterisation of a dna ligase gene of the extremely thermophilic archaeon desulfurolobus ambivalens shows close phylogenetic relationship to eukaryotic ligases. | a 3382 bp fragment containing a gene for a dna ligase from the extremely thermophilic, acidophilic, and facultatively anaerobic archaeon (archaebacterium) desulfurolobus ambivalens was cloned and sequenced. the deduced amino acid sequence (600 amino acids, 67619 molecular weight) showed 30-34% sequence identity with the atp-dependent eucaryal (eukaryotic) dna ligases of schizosaccharomyces pombe, saccharomyces cerevisiae, the human dna ligase i, and with the vaccinia dna ligase. distant similari ... | 1992 | 1437556 |
| domain structure of human 72-kda gelatinase/type iv collagenase. characterization of proteolytic activity and identification of the tissue inhibitor of metalloproteinase-2 (timp-2) binding regions. | the 72-kda gelatinase/type iv collagenase, a metalloproteinase thought to play a role in metastasis and in angiogenesis, forms a noncovalent stoichiometric complex with the tissue inhibitor of metalloproteinase-2 (timp-2), a potent inhibitor of enzyme activity. to define the regions of the 72-kda gelatinase responsible for timp-2 binding, a series of nh2- and cooh-terminal deletions of the enzyme were constructed using the polymerase chain reaction technique. the full-length and the truncated en ... | 1992 | 1322396 |
| infectious defective interfering particles of vsv from transcripts of a cdna clone. | the generation of infectious defective interfering (di) particles of vesicular stomatitis virus (vsv) entirely from cdna clones is reported. bacteriophage t7 rna polymerase was used to direct the transcription of a complete negative-stranded genomic rna from a cdna clone of a vsv di rna in cells simultaneously expressing the five vsv proteins from separately transfected cdna clones. the negative-stranded transcript was encapsidated with n protein, replicated by the vsv polymerase, and the replic ... | 1992 | 1318785 |
| functional interactions between the fusion protein and hemagglutinin-neuraminidase of human parainfluenza viruses. | the fusion glycoprotein (f) and hemagglutinin-neuraminidase (hn) genes of human parainfluenza virus type 2 (pi2) were molecularly cloned and expressed in hela-t4 cells by using the vaccinia virus-t7 transient expression system. expression of the f and hn proteins was detected by using immunoprecipitation and surface immunofluorescence staining. although the f protein was found to be cleaved into f1 and f2 and expressed on cell surfaces, no cell fusion was observed. however, cotransfection of the ... | 1992 | 1310764 |
| a plasmid that improves the efficiency of foreign gene expression by intracellular t7 rna polymerase. | to facilitate the construction of recombinant plasmids for expressing cloned genes with t7 rna polymerase supplied by recombinant vaccinia virus, a plasmid expression vector was designed by combining parts of plasmids ptz18r, pbluescript ii ks+, and par2529. the 3043-bp plasmid ptf1 has a t7 rna polymerase promoter, multiple cloning site for insertion of foreign genes, and a t7-specific transcription termination signal. plasmid ptf1 had several advantages compared with the reference plasmid par2 ... | 1992 | 1476796 |
| infectious japanese encephalitis virus rna can be synthesized from in vitro-ligated cdna templates. | japanese encephalitis virus (jev) is a positive-stranded enveloped rna virus that belongs to the family flaviviridae. genomic jev rna is approximately 11 kb long and encodes 10 proteins, 3 structural and 7 nonstructural. a full-length cdna copy of the jev genome was constructed by in vitro ligation of two cdna fragments which encode the 5' (nucleotide positions 1 to 5576) and 3' (nucleotide positions 5577 to 10976) halves of the genome. t7 rna polymerase transcripts of the ligated full-length cd ... | 1992 | 1501281 |
| studies on the initiation of simian virus 40 replication in vitro: rna primer synthesis and its elongation. | dna primase-dependent synthesis of oligoribonucleotides 10-15 nucleotides long was observed in the presence of atp, utp, gtp, and ctp by using the purified components of the simian virus 40 (sv40) dna replication system. the dna primase-catalyzed reaction required the sv40 large tumor antigen (t antigen), dna polymerase alpha (pol-alpha), the three-subunit human single-stranded dna binding protein (hssb), and topoisomerase i. the synthesis of small rnas was unaffected by the addition of activato ... | 1992 | 1310541 |
| cloning of the vaccinia virus ribonucleotide reductase small subunit gene. characterization of the gene product expressed in escherichia coli. | during its infectious cycle, vaccinia virus expresses a virus-encoded ribonucleotide reductase which is distinct from the host cellular enzyme (slabaugh, m.b., and mathews, c.k. (1984) j. virol. 52, 501-506; slabaugh, m.b., johnson, t.l., and mathews, c.k. (1984) j. virol. 52, 507-514). we have cloned the gene for the small subunit of vaccinia virus ribonucleotide reductase (designated vvr2) into escherichia coli and expressed the protein using a t7 rna polymerase plasmid expression system. afte ... | 1992 | 1309792 |
| the site-specific deoxyribonuclease from bacillus pumilus (endonuclease r.bpu1387). | a new site-specific endonuclease (dnase) was isolated from the cells of bacillus pumilus ahu 1387 strain. this enzyme (endonuclease r.bpu 1387) introduced double-stranded scissions at unique sites on dna's of coli phage lambda, lambdadvl, coli phage t7, bacillus phage phi105c, bacillus phage sp10, and simian virus 40, in the presence of magnesium ion. the activity was stimulated by the presence of nacl. | 1976 | 1018024 |
| e. coli membranes become permeable to ions following t7-virus-infection. | infection of e. coli with the viruses t7 or t3 leads to a dramatic efflux of potassium ions. this ion efflux is caused by the virus particle since no concomitant protein synthesis is required. t7 mutants carrying deletions in the m-gene (schweiger et al., 1975), however, yield virus particles disturbed in the ion release. | 1976 | 796675 |
| recombinant human immunodeficiency virus type 1 nucleocapsid (ncp7) protein unwinds trna. | the nucleocapsid protein (nc) of all animal retroviruses, encoded by the gag gene, is the major structural protein of the core ribonucleoprotein complex, bound to genomic rna in mature virions. nc is also thought to play one or more accessory roles in reverse transcription. mature nc (p7nc) from human immunodeficiency virus type 1 (hiv-1) is a 71-amino acid, basic protein which contains two cys3his zn(ii) retroviral-type zinc finger domains. herein, we describe the subcloning and expression of h ... | 1992 | 1551877 |
| physicochomecial studies on interactions between dna and rna polymerase. isolation and mapping of a t7 dna fragment containing the early promoters for escherichia coli rna polymerase. | the cleavage sites in the early promoter region of coliphage t7 have been mapped for four restriction enzymes. they are, from the left end in base pairs, 1100 and 740 for hinf; 680, 320, 530, 240, 77, and 67 for hind ii; 620 and 530 for hpa ii; 790 for alu i. the nucleotide sequence between the hind ii site at 680 base pairs from the left end and the hinf site at 740 base pairs from the left end has been determined, from which the start point of the promoter a3 is located at 720 base pairs from ... | 1976 | 795461 |
| 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 ... | 1992 | 1583735 |
| infectious in vitro transcripts from amplified cdnas of the y and kin strains of cucumber mosaic virus. | using a method based on the polymerase chain reaction (pcr) with primers that include the phi 10 promoter of bacteriophage t7, we obtained cdna clones of the three rna genomes of two different strains of cucumber mosaic virus (cmv; kin and y strains) from which infectious in vitro transcripts were generated, and demonstrated that the same primers could be used for amplification of at least two other strains of cmv (o and py). this method is rapid and requires only limited nucleotide (nt) sequenc ... | 1992 | 1601304 |
| filtration sizes of human immunodeficiency virus type 1 and surrogate viruses used to test barrier materials. | filters with well-defined holes were used to determine the effective diameters in buffer of human immunodeficiency virus type 1, herpes simplex virus type 1, and four bacteriophages (phi x174, t7, prd1, and phi 6), which may serve as surrogate viruses for testing barrier materials. bacteriophages phi 6 and prd1 most closely model human immunodeficiency virus type 1 in filtration size. | 1992 | 1610199 |
| spike protein-nucleocapsid interactions drive the budding of alphaviruses. | semliki forest virus (sfv) particles are released from infected cells by budding of nucleocapsids through plasma membrane regions that are modified by virus spike proteins. the budding process was studied with recombinant sfv genomes which lacked the nucleocapsid protein gene or, alternatively, the spike genes. no subviral particles were released from cells which expressed only the nucleocapsid protein or the spike proteins. virus release was found to be strictly dependent on the coexpression of ... | 1992 | 1629953 |
| bovine respiratory syncytial virus nucleocapsid protein: mrna sequence analysis and expression from recombinant vaccinia virus vectors. | the nucleotide sequence of the mrna encoding the nucleocapsid (n) protein of bovine respiratory syncytial (brs) virus, strain 391-2, was determined. recombinant vectors containing a cdna of the complete n gene were constructed, and expression of the n protein in eukaryotic cells was demonstrated using two different vector systems. the brs virus n mrna was 1197 nucleotides in length, exclusive of poly(a), and had a single major open reading frame that encoded a polypeptide of 391 amino acids with ... | 1992 | 1634882 |
| comparison of biotinylated dna and rna probes for rapid detection of varicella-zoster virus genome by in situ hybridization. | we describe a general method for the production of nonisotopic dna and rna probes for the detection of the varicella-zoster virus (vzv) genome by in situ hybridization. vzv dna was extracted from purified viral nucleocapsids, cleaved with restriction enzyme (re) bamhi, and cloned into plasmid pbr322 by the standard vector insert procedure. we cloned over 85% of the vzv genome and obtained 18 recombinants. plasmids containing the b, f, g, h, and j fragments of vzv dna were labeled by the nick tra ... | 1991 | 1645371 |
| overexpression in bacterial and identification in infected cells of the pseudorabies virus protein homologous to herpes simplex virus type 1 icp18.5. | the icp18.5 gene (ul28) of herpes simplex virus type 1 is a member of a well-conserved gene family among herpesviruses and is thought to play a role in localization of viral glycoproteins. we have cloned, sequenced, and expressed the entire pseudorabies virus (prv) icp18.5 open reading frame in escherichia coli as a cro-icp18.5 fusion protein. rabbit antiserum against cro-icp18.5 immunoprecipitated a 79-kda protein from prv-infected cells as well as a 79-kda protein from in vitro translation of ... | 1991 | 1645790 |
| membrane alteration induced by t7 virus infection. | | 1977 | 320041 |
| purification and characterization of the herpes simplex virus type 1 ribonucleotide reductase small subunit following expression in escherichia coli. | the herpes simplex virus type 1 (hsv-1) gene encoding the ribonucleotide reductase (rr) small subunit (r2) was cloned as an unfused and intact open reading frame into a t7 rna polymerase expression system in escherichia coli. the expressed product was recovered from bacteria in soluble form and constituted 7% of the soluble protein. protein purification yielded 3.5 mg of 95% pure r2 per litre of bacterial culture. the correct composition of the purified protein was verified by amino acid analysi ... | 1991 | 1646278 |
| adenovirus dna replication in vitro. | a soluble extract from the nuclei of hela cells infected with adenovirus 5 (ad5) carries out the semiconservative replication of exogenously added ad5 dna in vitro. maximal dna synthesis is observed when dna-protein complex, isolated from ad5 virions, is added as template. dna-protein complex from virions of the closely related virus, adenovirus 2, is also active. in contrast, very little in vitro dna synthesis is observed when deproteinized ad5 dna or dna from a heterologous source (bacteriopha ... | 1979 | 284391 |
| direct introduction and transient expression of capped and non-capped rna in saccharomyces cerevisiae. | we report the introduction of functional rna molecules into yeast spheroplasts. plasmids containing the firefly luciferase coding region were transcribed to yield rnas suitable for introduction into yeast cells and direct assay of their translation products. the 5' noncoding regions of the rnas were derived either from the 5' noncoding regions of firefly luciferase, poliovirus, or yeast virus-like-particle (vlp) l-a or m1 rnas. capped and non-capped mrnas were made by t7 rna polymerase-directed ... | 1991 | 1656383 |
| involvement of eucaryotic deoxyribonucleic acid polymerases alpha and gamma in the replication of cellular and viral deoxyribonucleic acid. | in an effort to identify the deoxyribonucleic acid (dna) polymerase activities responsible for mammalian viral and cellular dna replication, the effect of dna synthesis inhibitors on isolated dna polymerases was compared with their effects on viral and cellular dna replication in vitro. dna polymerase alpha, simian virus 40 (sv40) dna replication in nuclear extracts, and cv-1 cell (the host for sv40) dna replication in isolated nuclei all responded to dna synthesis inhibitors in a quantitatively ... | 1979 | 226127 |
| an electron microscopic method for the mapping of proteins attached to nucleic acids. | an electron microscopic method for demonstrating the presence of and mapping the positions of proteins specifically bound to nucleic acids is described. the nucleic acid-protein complex is treated with dinitrofluorobenzene under conditions such that dinitrophenyl (dnp) groups are attached to nucleophilic groups on the protein, with only a low level of random attachment to the nuclei acid. this product is treated with rabbit anti-dnp igg. the position of the protein-(dnp)n(igg)m complex on the nu ... | 1978 | 211486 |
| requirement of the bacteriopahge t7 0.7 gene for phage growth in the presence of the col 1b factor. | a variety of colicinogenic factors were examined for their effect on the growth of phages related to t7. col b1 and col b4 inhibited the growth of every phage tested. col b2, however, did not interfere with t3 and h. col e2 interfered only with growth of t3, while col ib produced abortive infection with some t7 mutants. the abortive infection associated with col ib seems to be correlated with the absence of the virus phosphokinase; mixed infection with t7 wild type and a phosphokinase deficient ... | 1977 | 192846 |
| enhancement of the vaccinia virus/phage t7 rna polymerase expression system using encephalomyocarditis virus 5'-untranslated region sequences. | a recombinant vaccinia virus producing the bacteriophage t7 rna polymerase was used to express foreign genes in eukaryotic cells. translation efficiency in this expression system was enhanced significantly by employing the encephalomyocarditis virus (emcv) 5'-untranslated region (utr) which confers cap-independent translation by directing internal initiation of translation. the enhancement was accomplished by fusing open reading frames (orfs) to the n terminus of the emcv polyprotein coding regi ... | 1991 | 1660838 |
| high-efficiency protein synthesis from t7 rna polymerase transcripts in 3t3 fibroblasts. | after nih3t3 cells constitutively expressing t7 rna polymerase were transfected (+ ca.phosphate) with a circular dna containing the firefly luciferase(luc)-encoding gene (luc) 3' to the encephalomyocarditis (emc) virus 5'-untranslated sequence and t7 promoter, luc protein comprising approx. 20% of total cellular protein was obtained. after similar transfection of an analogous construct containing the lacz gene into the same cell line, at least 50% of the cells produced beta-galactosidase. fibrob ... | 1991 | 1662654 |
| partial purification and properties of a dna-binding protein from nuclei of cells infected with polyoma virus. | a dna-binding protein has been purified from nuclei of 3t3 cells infected with polyoma virus. the assay used to detect this activity measures the amount of double-stranded dna retained on a nitrocellulose membrane filter in the presence of binding protein. the interaction between dna and protein is salt dependent and occurs optimally at 0.8 m nacl. the isolated protein can bind to both circular and linear duplex dna. incubation of the binding protein with pm2 or polyoma dna results in the format ... | 1976 | 174725 |
| infectious rna transcripts from ross river virus cdna clones and the construction and characterization of defined chimeras with sindbis virus. | we have constructed a full-length cdna clone of the virulent t48 strain of ross river virus, a member of the alphavirus genus. infectious rna can be transcribed from this clone using sp6 or t7 rna polymerase. the rescued virus has properties indistinguishable from those of the t48 strain of ross river virus. we have used this clone, together with a full-length cdna clone of sindbis virus, to construct chimeric plasmids in which the 5' and the 3' nontranslated regions of the sindbis and ross rive ... | 1991 | 1673812 |
| synthesis of infectious rna from full-length cloned cdna to rna of cymbidium ringspot tombusvirus. | a full-length dna copy of cymbidium ringspot virus rna was cloned downstream of a phage t7 promoter. in vitro transcripts had no extra nucleotides at the 3' terminus, and a 5' end likely to be precisely as in genomic rna. transcripts were infective when inoculated into test plants. northern blots from inoculated plants revealed the presence of genomic and subgenomic rnas, but not of satellite rna. virus particles isolated from infected plants had the same outward aspect and size as those of the ... | 1990 | 1697330 |
| inhibition of expression of human immunodeficiency virus-1 in vitro by antibody-targeted liposomes containing antisense rna to the env region. | previous studies revealed that antisense oligodeoxynucleotides to specific regions of the human immunodeficiency virus-1 (hiv-1) are potent inhibitors of replication of hiv-1 in vitro (zamecnik, p. c., goodchild, j., taguchi, y., and sarin, p. s. (1986) proc. natl. acad. sci. u. s. a. 83, 4143-4146). we now report that antisense rna, synthesized in vitro using t7 and sp6 rna polymerase, displayed an anti-hiv-1 effect in the htlv-iiib/h9 system in vitro. treatment of hiv-1-infected h9 cells with ... | 1990 | 1697856 |
| [effective synthesis of oligo(poly)deoxyribonucleotides using an h-phosphonate method in plastic microcolumn]. | a facile technique of manual oligonucleotide synthesis via h-phosphonate approach is developed. syntheses carried out in pipette tips with siliconised glasswool filters take 3-3.5 min per cycle with 97-98% yields per condensation. the method was used to synthesize 12-55-mers: t7 and pl promoter regions, gene of the signal peptide of the e. coli ompa protein, oligonucleotides coding for amino acid sequences 94-105 of pres1- and 133-143 of pres2-regions of hepatitis b virus, hybridisation probes, ... | 1990 | 1700717 |
| effects of 2-chloro-2'-deoxyadenosine 5'-triphosphate on dna synthesis in vitro by purified bacterial and viral dna polymerases. | 2-chloro-2'-deoxyadenosine 5'-triphosphate (cldatp) was compared with datp as a substrate for dna synthesis by bacterial and viral dna polymerases in vitro. lengths of chain extension and dna synthesis pause sites were determined by comparison with products generated by dideoxynucleotide sequencing methods on the same end-labeled primer/template duplex after high-resolution polyacrylamide gel electrophoresis. reverse transcriptase (rt) from human immunodeficiency virus (hiv-1) and avian myelobla ... | 1991 | 1703019 |
| bacteriophage t5-induced endonucleases that introduce site-specific single-chain interruptions in duplex dna. | four site-specific endodeoxyribonucleases have been partially purified from extracts of bacteriophage t5-infected escherichia coli by gel filtration and affinity chromatography on single- and double-stranded dna. the enzymes were detected and characterized by agarose gel electrophoresis of alkali-denatured digestion products. none of the four is found in uninfected cells. in the presence of a divalent cation, all four endonucleases make ligase-repairable, single-chain interruptions at specific s ... | 1976 | 5725 |
| gtp-binding mutants of rab1 and rab2 are potent inhibitors of vesicular transport from the endoplasmic reticulum to the golgi complex. | we have examined the role of ras-related rab proteins in transport from the er to the golgi complex in vivo using a vaccinia recombinant t7 rna polymerase virus to express site-directed rab mutants. these mutations are within highly conserved domains involved in guanine nucleotide binding and hydrolysis found in ras and all members of the ras superfamily. substitutions in the gtp-binding domains of rab1a and rab1b (equivalent to the ras 17n and 116i mutants) resulted in proteins which were poten ... | 1992 | 1429835 |
| translational modulation in hepatitis b virus pres-s open reading frame expression. | a series of hepatitis b virus open reading frame (orf) pres-s variants, including mutants in which the relative order of the in-frame start codons (aug1, aug2 and aug3) and nearby sequences had been altered, was expressed both in vivo (in hepg2 hepatoblastoma cells) and in vitro (by t7 promoter-driven transcription followed by translation in a rabbit reticulocyte lysate). the ratio of the synthesis of the large, middle (m) and major (s) proteins or their chimeric counterparts was analysed to stu ... | 1992 | 1730934 |
| the role of conserved aspartate and serine residues in ligand binding and in function of the 5-ht1a receptor: a site-directed mutation study. | wild-type and mutant serotonin 1a receptors were transiently expressed in cos-7 cells using the infection-transfection variant of the vaccinia virus/t7 polymerase vector system. the amino acid substitutions in the transmembrane regions, asp82-->asn82, asp116-->asn116, and ser198-->ala198 all resulted in a decrease in affinity for 5-ht by 60-100-fold, without affecting the affinity for the antagonist, pindolol. the binding of agonist to the additional mutant, thr199-->ala199, was too weak to be m ... | 1992 | 1426261 |
| mutagenesis of the l protein encoded by bunyamwera virus and production of monospecific antibodies. | bacterial fusion proteins containing portions of the bunyamwera virus l protein were used as immunogens to prepare antisera in rabbits. of five fusion proteins injected into rabbits, three yielded sera that reacted with the bunyamwera virus l protein, detected by western blotting or immunoprecipitation. two of these antisera were specific for either the amino- or carboxy-terminal regions of the l protein. the specificity of these antisera was confirmed by their pattern of reactivity with full-le ... | 1992 | 1402814 |
| specific valylation of turnip yellow mosaic virus rna by wheat germ valyl-trna synthetase determined by three anticodon loop nucleotides. | the valylation by wheat germ valyl-trna synthetase of anticodon loop mutants of turnip yellow mosaic virus rna has been studied. rna substrates 264 nucleotides long were made by t7 rna polymerase from cdna encompassing the 3' trna-like region of genomic rna. substitution singly, or in combination, of three nucleotides in the anticodon loop resulted in very poor valylation (vmax/km less than 10(-3) relative to wild type). these nucleotides thus represent the major valine identity determinants rec ... | 1992 | 1390705 |
| dna synthesis blocking lesions induced by singlet oxygen are targeted to deoxyguanosines. | in vitro dna synthesis on single stranded templates damaged by singlet oxygen was investigated in the supf trna gene sequence, using several dna polymerases. singlet oxygen was generated by the thermal decomposition of the water soluble with the endoperoxide of disodium 3,3'-(1,4-naphthylidene) dipropionate (ndpo2). the data demonstrated that damage at deoxyguanosine residues interrupts dna polymerization. modified t7 phage and thermus aquaticus dna polymerases were found to synthesize dna fragm ... | 1992 | 1375992 |
| renal and adrenal sympathetic preganglionic neurons in rabbit spinal cord: tracing with herpes simplex virus. | we mapped sympathetic renal preganglionic neurons in the rabbit spinal cord using herpes simplex virus retrograde transneuronal tracing after application of the virus to the renal nerve. virus-positive neurons were found in the spinal cord from t7 to l2, principally in the ipsilateral intermediolateral cell column. renal and adrenal preganglionic neurons are largely separate populations. extensive nonspecific spread of virus from infected cells to neighboring neurons does not appear to occur. | 1992 | 1374281 |
| cells that express all five proteins of vesicular stomatitis virus from cloned cdnas support replication, assembly, and budding of defective interfering particles. | an alternative approach to structure-function analysis of vesicular stomatitis virus (vsv) gene products and their interactions with one another during each phase of the viral life cycle is described. we showed previously by using the vaccinia virus-t7 rna polymerase expression system that when cells expressing the nucleocapsid protein (n), the phosphoprotein (ns), and the large polymerase protein (l) of vsv were superinfected with defective interfering (di) particles, rapid and efficient replic ... | 1991 | 1847519 |
| dna-binding activity of the murine homeodomain protein hox-2.3 produced by a hybrid phage t7/vaccinia virus system. | homeobox-containing genes encode transcription factors that, via the homeodomain, bind specifically to dna. to study the dna-binding properties of the murine homeodomain-containing protein, hox-2.3, a hybrid expression system was used, combining gene expression by recombinant vaccinia virus (revv) with bacteriophage t7 transcription. expression was achieved by co-infecting hela cells with two revvs, one expressing the t7-rna polymerase-encoding gene directed by the vv promoter, p7.5, and another ... | 1992 | 1353046 |
| direct double-stranded dna sequencing with baculovirus genomes. | double-stranded dna sequencing with the modified t7 dna polymerase (sequenase) was performed directly with nuclear polyhedrosis virus dna genomes. the conditions were optimized to allow for a rapid and unambiguous sequence analysis of nuclear polyhedrosis virus genomes. | 1991 | 1849913 |
| detection of hepatitis a virus and other enteroviruses in water by ssrna probes. | sensitive and specific methods are needed to detect hepatitis a virus (hav) and other human enteroviruses in environmental samples such as drinking water and foods. clones of cdna encoding the 5'-most 1 kb of the hav and coxsackievirus b3 (cb3) genomes were subcloned into t7/sp6 rna transcription vectors. in vitro transcribed rna from the t7 promoter detected their respective hav or cb3 genomic rna. conversely, sp6 transcripts detected viral negative-stranded rna but not the genome. when both ss ... | 1991 | 1849914 |
| trans rescue of a mutant poliovirus rna polymerase function. | a series of three-nucleotide insertions was engineered into the p2 and p3 coding regions of the t7 expression plasmid pt7(tau)-pv1, which encodes a full-length copy of poliovirus type 1 (mahoney) cdna. when rna derived in vitro from these mutated templates was used to transfect hela cells, viable virus mutants were recovered. one mutant, sel-3d-18, which contained a single amino acid insertion in the 3dpol coding region, was temperature sensitive for growth at 39 degrees c and showed defects in ... | 1991 | 1850039 |
| identification and functional characterization of epstein-barr virus dna polymerase by in vitro transcription-translation of a cloned gene. | in order to identify the gene encoding the epstein-barr virus (ebv) dna polymerase, a portion of the bamhi-a fragment containing the fifth leftward open reading frame (balf5) of the ebv genome was cloned into sp6 and t7 promoter-containing vectors for in vitro transcription-translation. the rna synthesized in vitro was used to program rabbit reticulocyte lysates, which were analyzed for the synthesis of the putative polymerase polypeptide (110 kda) and assayed directly for ebv dna polymerase act ... | 1991 | 1850046 |
| the 5'-terminal nucleotides of hepatitis a virus rna, but not poliovirus rna, are required for infectivity. | a series of plasmids containing hepatitis a virus (hav) cdna was constructed such that positive-strand hav rna could be transcribed with t7 rna polymerase. the plasmids differed in the number of 5'-terminal nucleotides representing the junctions between vectors and hav sequences that were present in the transcripts. when these transcripts were used to transfect cultured bs-c-1 cells, it was found that only those transcripts that contained all of the 5'-terminal hav nucleotides, in addition to on ... | 1991 | 1850050 |
| the large subunit of herpes simplex virus type 1 ribonucleotide reductase: expression in escherichia coli and purification. | the open reading frame of the large subunit (r1) of herpes simplex virus type 1 (hsv-1) ribonucleotide reductase has been positioned downstream of the phage t7 gene 10 promoter in the expression vector, pet. transformation of this recombinant plasmid into escherichia coli bl21 de3 cells containing the t7 rna polymerase, under the control of the lac uv5 promoter, allows expression of the subunit on induction of the t7 rna polymerase by isopropyl thiodigalactoside. the expressed protein is soluble ... | 1991 | 1850930 |
| in vitro translation of hepatitis a virus subgenomic rna transcripts. | a subgenomic cdna clone from hepatitis a virus strain hm175, composed of the last eight nucleotides of the 5' non-translated region and the first 2248 nucleotides of the coding sequence (p1 region), was inserted into a vector under the control of the t7 promoter. restriction enzyme digestion at sites within the structural region and subsequent transcription in vitro yielded rna products which were translated efficiently in rabbit reticulocyte lysates to produce proteins of the predicted sizes. t ... | 1991 | 1851807 |
| expression of human immunodeficiency virus type 1 (hiv-1) gag, pol, and env proteins from chimeric hiv-1-poliovirus minireplicons. | recent studies have demonstrated that genomes of poliovirus with deletions in the p1 (capsid) region contain the necessary viral information for rna replication. to test the effects of the substitution of foreign genes on rna replication and protein expression, chimeric human immunodeficiency virus type 1 (hiv-1)-poliovirus genomes were constructed in which regions of the gag, pol, or env gene of hiv-1 were substituted for regions of the p1 gene in the infectious cdna clone of type 1 mahoney pol ... | 1991 | 1851859 |
| in vitro membrane binding of the translation products of the carlavirus 7-kda protein genes. | two double-stranded dna copies of the genes potentially coding for the 7-kda proteins of potato virus m (pvm) and potato virus s (pvs) were synthesized and cloned into t7 transcription vectors. cell-free translation of the corresponding monocistronic transcripts yielded in both cases a single protein of approximately 7-8 kda that contains a highly hydrophobic n-terminal segment. to analyze their membrane-binding potential, both proteins were synthesized in the membrane-enriched krebs-2 extract. ... | 1991 | 1853576 |
| a chimeric avian retrovirus containing the influenza virus hemagglutinin gene has an expanded host range. | we have investigated what protein sequences are necessary for glycoprotein incorporation into rous sarcoma virus (rsv) virions by utilizing the hemagglutinin (ha) protein of influenza virus. two chimeric ha genes were constructed. in the first the coding sequence for the signal peptide of the rsv env gene product was fused in frame to the entire ha structural gene, and in the second the hydrophobic anchor and cytoplasmic domain sequences of the ha gene were also replaced with those from the rsv ... | 1992 | 1331528 |
| purification and characterization of human papillomavirus type 16 e7 protein with preferential binding capacity to the underphosphorylated form of retinoblastoma gene product. | human papillomavirus type 16 e7 is considered to be a major viral oncoprotein playing an important role(s) in cervical cancers. e7 protein was shown to bind to the protein product of the retinoblastoma gene (rb), while simian virus 40 large t and adenovirus e1a were also shown to possess binding activity to rb protein. the rb protein is a cell cycle regulator that is highly phosphorylated specifically in s, g2, and m, whereas it is underphosphorylated in g0 and g1. recently, large t was demonstr ... | 1991 | 1870208 |
| cloned dna copies of cowpea severe mosaic virus genomic rnas: infectious transcripts and complete nucleotide sequence of rna 1. | cowpea severe mosaic virus (cpsmv) is a member of the comovirus group of messenger-sense rna viruses with bipartite genomes, of which cowpea mosaic virus (cpmv) is the type member. full-length copies of cpsmv rna 1 were cloned in plasmids bearing a bacteriophage t7 promoter. previously, similar clones of cpsmv rna 2 had been obtained. a 5'-ruauuaaaauuuu sequence is common to rna 1 and rna 2. from two rna 1 clones and four rna 2 clones we excised non-cpsmv sequences so as to provide templates for ... | 1992 | 1448917 |
| overproduction of adenovirus dna polymerase and preterminal protein in hela cells. | adenovirus (ad) dna polymerase (adpol) and the preterminal protein (ptp) form a complex that is involved in the in vitro initiation of ad dna replication. recombinant vaccinia viruses (vv) were constructed in which the genes encoding adpol and ptp were cloned into a vaccinia/t7 hybrid expression-based vector downstream from the t7 promoter (pt7)/encephalomyocarditis virus (emcv) 5'-untranslated region (utr). hela cells infected with the recombinant vv-adpol or vv-ptp or a mixture of both, togeth ... | 1991 | 1937014 |
| infectious in vitro rna transcripts derived from cloned cdna of the cucurbit potyvirus, zucchini yellow mosaic virus. | a full-length cdna clone of the rna genome of the cucurbit potyvirus zucchini yellow mosaic virus (zymv) was constructed downstream from a bacteriophage t7 rna polymerase promoter. a single extra guanosine residue not present in zymv rna was added to the 5' and 3' ends. capped (m7gpppg) zymv rna transcripts were infectious in 10 of 91 cucurbita pepo test plants; uncapped rna transcripts were not infectious. the appearance of symptoms in plants inoculated with the infectious transcript was delaye ... | 1991 | 1940860 |
| synthesis and localization of japanese encephalitis virus rnas in the infected cells. | synthesis and localization of virus-specific rna in cells infected with japanese encephalitis virus (jev) were examined. to prepare specific rna probes, we constructed four kinds of plasmids which contained dna fragments corresponding to jev genomic rna. minus probes, jt18v and jt19iii, transcribed by t7 rna polymerase were able to recognize a negative strand of jev-specific rna synthesized in cells as early as 6 hr postinfection (p.i.). in the experiments using a plus-strand probe jt19v to hybr ... | 1990 | 1963921 |
| expression of bluetongue virus serotype 17 ns1 protein from a cloned gene. | a full-length copy of the coding region of segment 6 from bluetongue virus (btv) serotype 17 was constructed from five overlapping cdna clones. the gene coding for the ns1 protein was cloned into an expression plasmid under the control of a bacteriophage t7 promoter and expressed both in vitro and in escherichia coli bl21(de3) cells which contain a t7 rna polymerase gene in their chromosome. expression in both systems resulted in the synthesis of a protein comigrating with ns1 and a minor polype ... | 1990 | 1964519 |
| encoding of a homolog of the ifn-gamma receptor by myxoma virus. | many poxvirus-encoded virulence factors have been identified as proteins that are secreted from infected cells. the major secreted protein (37 kilodaltons) from cells infected with myxoma virus is encoded by the m-t7 open reading frame. this protein has significant sequence similarity to the human and mouse receptors for interferon-gamma (ifn-gamma). furthermore, the myxoma m-t7 protein specifically binds rabbit ifn-gamma and inhibits the biological activity of extracellular ifn-gamma, one of th ... | 1992 | 1455233 |
| pseudouridine in the anticodon g psi a of plant cytoplasmic trna(tyr) is required for uag and uaa suppression in the tmv-specific context. | we have previously isolated and sequenced nicotiana cytoplasmic trna(tyr) with g psi a anticodon which promotes readthrough over the leaky uag termination codon at the end of the 126 k cistron of tobacco mosaic virus rna and we have demonstrated that trna(tyr) with q psi a anticodon is no uag suppressor. here we show that the nucleotide in the middle of the anticodon (i.e., psi 35) also contributes to the suppressor efficiency displayed by cytoplasmic trna(tyr). a trna(tyr) with gua anticodon wa ... | 1992 | 1461724 |
| expression in animal cells of the 5-ht1a receptor by a vaccinia virus vector system. | the co-infection or infection-transfection variants of the t7 rna polymerase/vaccinia vector system were used to express 5-ht1ars in cos-7, bsc-40 and gh3 cells, with co-infection giving ca. 3-fold higher level than infection-transfection. binding affinities were similar to those of the endogenous 5-ht1ar, with highest affinities for 5-ht and 8-oh-dpat. functional properties were demonstrated by assays of agonist-stimulated gtpase activity and its inhibition by pertussin toxin. immunoblot assays ... | 1992 | 1533596 |
| association of the nonstructural protein nss of uukuniemi virus with the 40s ribosomal subunit. | the small rna segment (s segment) of uukuniemi (uuk) virus encodes two proteins, the nucleocapsid protein (n) and a nonstructural protein (nss), by an ambisense strategy. the function of nss has not been elucidated for any of the bunyaviruses expressing this protein. we have now expressed the n and nss proteins in sf9 insect cells by using the baculovirus expression system. high yields of both proteins were obtained. a monospecific antibody was raised against gel-purified nss and used to study t ... | 1992 | 1534850 |
| expression of a cloned gamma-aminobutyric acid transporter in mammalian cells. | the cdna clone gat-1, which encodes a na(+)- and cl(-)-coupled gaba transporter from rat brain, has been expressed in mammalian cells using three different systems: (1) transient expression upon transfection of mouse ltk- cells with a eukaryotic expression vector containing gat-1; (2) stable expression in l-cells transfected with the same vector; (3) transfection of hela cells infected with a recombinant vaccinia virus expressing t7 rna polymerase. similar results both qualitatively and quantita ... | 1992 | 1536839 |
| cell-specific posttranslational events affect functional expression at the plasma membrane but not tetrodotoxin sensitivity of the rat brain iia sodium channel alpha-subunit expressed in mammalian cells. | the rat brain iia na+ channel alpha-subunit was expressed and studied in mammalian cells. cells were infected with a recombinant vaccinia virus (vv) carrying the bacteriophage t7 rna polymerase gene and were transfected with cdna encoding the iia na+ channel alpha-subunit under control of a t7 promoter. whole-cell patch-clamp recording showed that functional iia channels were expressed efficiently (approximately 10 channels/microns2 in approximately 60% of cells) in chinese hamster ovary (cho) c ... | 1992 | 1309573 |
| cellular expression of a functional nodavirus rna replicon from vaccinia virus vectors. | rna replication provides a powerful means for the amplification of rna, but to date it has been found to occur naturally only among rna viruses. in an attempt to harness this process for the amplification of heterologous mrnas, both an rna replicase and its corresponding rna templates have been expressed in functional form, using vaccinia virus-bacteriophage t7 rna polymerase vectors. plasmids were constructed which contained in 5'-to-3' order (i) a bacteriophage t7 promoter; (ii) a full-length ... | 1992 | 1548766 |
| a quantifiable phenotype of viral propagation. | a system has been identified where a virus, replicating continuously on its host, displays a distinct and quantifiable phenotype, and thereby continuously reports on the state of the virus-host relationship. when bacteriophage t7 is plated out with its host, escherichia coli, it establishes a constant-velocity infection wave, which is driven by an autocatalytic reaction-diffusion mechanism. the velocity--which is easily measured--continuously reflects the infection environment. the simplicity of ... | 1991 | 1993042 |
| in vivo complementation of infectious transcripts from mutant tobacco mosaic virus cdnas in transgenic plants. | a full-length cdna clone of the u1 (common) strain of tobacco mosaic virus (tmv) was constructed, and highly infectious transcripts were produced in vitro using bacteriophage t7 rna polymerase. frameshift mutations designed to cause premature termination of translation were introduced into either the 30-kda movement protein (mp) gene or the coat protein (cp) gene. the mp-frameshift mutant was unable to locally or systemically infect inoculated tobacco plants. however, inoculation of transgenic t ... | 1991 | 1994570 |