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immune responses to sivmne envelope glycoproteins protect macaques from homologous siv infection. 19921466988
phenotypic characterization of three temperature-sensitive mutations in the vaccinia virus early gene transcription initiation factor.vaccinia virus gene d6r encodes the small subunit of the virion early gene transcription initiation factor. three temperature-sensitive mutations have been mapped to this gene. the biochemical phenotype exhibited by each mutation was examined. all mutants displayed altered viral protein synthesis in pulse-labelling analyses at both the permissive and non-permissive temperatures. the onset of early protein synthesis was delayed, and the rate of early protein synthesis was reduced in each case. fu ...19921469354
comparison of the thymidine kinase genes from three entomopoxviruses.the entomopoxviruses (insect poxviruses) of eastern spruce budworm (choristoneura fumiferana), two year cycle spruce budworm (c. biennis) and the indian red army worm (amsacta moorei) are being studied in our laboratory for their potential as biological insecticides and expression vectors. these viruses characteristically replicate in the cytoplasm of insect cells and produce occlusion bodies that serve to protect the virion from the environment. by analogy to mammalian poxviruses, they should a ...19921469363
in vitro virucidal effects of allium sativum (garlic) extract and compounds.garlic (allium sativum) has been shown to have antiviral activity, but the compounds responsible have not been identified. using direct pre-infection incubation assays, we determined the in vitro virucidal effects of fresh garlic extract, its polar fraction, and the following garlic associated compounds: diallyl thiosulfinate (allicin), allyl methyl thiosulfinate, methyl allyl thiosulfinate, ajoene, alliin, deoxyalliin, diallyl disulfide, and diallyl trisulfide. activity was determined against s ...19921470664
targeting of sp1 to a non-sp1 site in the human neurofilament (h) promoter via an intermediary dna-binding protein.the human neurofilament (h) promoter contains multiple binding sites for nuclear proteins including a proximal (prox) site centered around the sequence ggttggacc and an adjacent pyrimidine (pyr) tract site centered around the sequence ccctcctcccc. surprisingly binding to a probe containing the prox/pyr region of the nf(h) promoter was competed in gel shifts by an oligonucleotide containing only an sp1 binding site (ggggcgggg). supershift assays with a polyclonal anti-sp1 antisera confirmed that ...19921475189
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 ...19921476796
in vitro infection of human placental trophoblast by wild-type vaccinia virus and recombinant virus expressing hiv envelope glycoprotein.short-time (< or = 7 days) cultures of trophoblast mononuclear cells isolated from term placentae were challenged with vaccinia virus. cytopathic effects were induced in crude placental cell preparations as well as in cultures established after negative immunosorting of major histocompatibility complex class i epitope-expressing cells, i.e. cultures exclusively derived from villous cytotrophoblast according to our present state of knowledge. the trophoblast in vitro supported a full replicative ...19921480824
antisense oligodeoxynucleotides as antiviral agents. 19921486989
localization of rab family members in animal cells. 19921488012
the n-terminal 22 amino acids encoded by the gene specifying the major secreted protein of vaccinia virus, strain lister, can function as a signal sequence to direct the export of a foreign protein.cells infected with vaccinia virus strain lister secrete a polypeptide of approximate molecular weight 35,000 (35k) into the medium. previous studies identified a cleavable, hydrophobic region of 17 amino acids in the 35k protein which could potentially function as a signal peptide to target the protein to the secretory pathway. here we report the use of the expression-secretion signals derived from the 35k gene to direct export and secretion of a foreign protein. vaccinia virus recombinants car ...19921492495
stable expression of the human immunodeficiency virus type 1 envelope glycoprotein in transfected l cells.an sv40-based expression vector was used to generate cd4-negative murine l cell lines which stably expressed the human immunodeficiency virus envelope glycoprotein (env). despite the presence of abundant intracellular envelope glycoprotein, the expression of env gp120/41 was not detected on the cell surface. pulse-chase studies showed that the majority of the gp120 detected at the end of a 20-h chase was in the culture medium. therefore gp120 was shed and/or secreted from these cells. transfecte ...19921493050
glutaredoxin homolog encoded by vaccinia virus is a virion-associated enzyme with thioltransferase and dehydroascorbate reductase activities.glutaredoxins (grxs), also known as thioltransferases, use glutathione as a cofactor for reduction of disulfides in prokaryotes and eukaryotes. we demonstrate that the vaccinia virus o2l open reading frame encodes a functional grx, as predicted by johnson et al. [johnson, g. p., goebel, s. j., perkus, m. e., davis, s. w., winslow, j. p. & paoletti, e. (1991) virology 181, 378-381] from sequence homology. the 12-kda protein product of the o2l open reading frame was synthesized after viral dna rep ...19921496000
vaccinia virus infection induces a stress response that leads to association of hsp70 with viral proteins.we studied the impact of vaccinia virus infection on stress protein gene expression in human cells and investigated the possibility that eukaryotic heat shock proteins interact with viral components during assembly. infection of human monocyte-macrophages by vaccinia virus caused a dramatic decrease in levels of cellular mrnas such as those encoding actin and tubulin. in contrast, infection did not cause a significant reduction in the levels of hsp90 and hsp60 mrnas and led to substantially incr ...19921501279
expression of the influenza a virus m2 protein is restricted to apical surfaces of polarized epithelial cells.the m2 protein of influenza a virus is a small, nonglycosylated transmembrane protein that is expressed on surfaces of virus-infected cells. a monoclonal antibody specific for the m2 protein was used to investigate its expression in polarized epithelial cells infected with influenza virus or a recombinant vaccinia virus that expresses m2. the expression of m2 on the surfaces of influenza virus-infected cells was found to be restricted to the apical surface, closely paralleling that of the influe ...19921501289
steroid hormone synthesis by a vaccinia enzyme: a new type of virus virulence factor. 19921505526
[identification of the gene for the immunodominant p35 protein from vaccinia virus].a major immunodominant envelope protein p35 of vaccina virus was purified by means of extraction from virions with detergent np-40. the protein was cleaved with cnbr, four homogenous peptides were isolated and their n-terminal amino acid sequences were determined. computer search in a protein sequences data bank revealed that the immunodominant protein p35 of vaccinia virus is encoded by h3 gene in hindiii-h fragment of vaccinia virus genome.19921508163
purification and characterization of dna ligase i from the trypanosomatid crithidia fasciculata.a dna ligase has been purified approximately 5000-fold, to near homogeneity, from the trypanosomatid crithidia fasciculata. the purified enzyme contains polypeptides with molecular masses of 84 and 80 kda as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. both polypeptides formed enzyme-adenylate complexes in the absence of dna, contained an epitope that is highly conserved between human and bovine dna ligase i and yeast and vaccinia virus dna ligases, and were identified ...19921508676
molecular cloning and expression of a novel keratinocyte protein (psoriasis-associated fatty acid-binding protein [pa-fabp]) that is highly up-regulated in psoriatic skin and that shares similarity to fatty acid-binding proteins.analysis by means of two-dimensional (2d) gel electrophoresis of the protein patterns of normal and psoriatic unfractionated non-cultured keratinocytes has revealed a few low-molecular-weight proteins that are highly up-regulated in psoriatic skin. these include psoriasin; calgranulin b, also known as mrp 14, l1, or calprotectin; calgranulin a or mrp 8; and cystatin a or stefin a. here, we have cloned and sequenced the cdna (clone 1592) encoding a new member of this group of low-molecular-weight ...19921512466
interactions of cd4+ plasma membrane vesicles with hiv-1 and hiv-1 envelope glycoprotein-expressing cells.to study interactions between the human immunodeficiency virus type 1 (hiv-1) envelope glycoprotein (gp120-gp41) and the receptor in the target membrane, cd4, a new experimental system utilizing cd4-carrying plasma membrane vesicles (cd4 pmvs) was developed. cd4 pmvs were prepared by hypotonic lysis of hela cells expressing cd4 after infection with recombinant vaccinia virus containing the cd4 cdna. the cd4 pmvs carried up to 680 cd4 molecules per vesicle. their fusion with cells expressing gp12 ...19921512692
biological effects of recombinant vaccinia virus-expressed interleukin 4.recombinant vaccinia viruses expressing murine interleukin 4 (il-4), either alone or together with interleukin 2 (il-2) or gamma interferon (gamma-ifn), were constructed. unlike il-2, il-4 expressing viruses were not cleared from immunodeficient mice and the mice died. as they died more rapidly than immunodeficient mice inoculated with a control virus, it appeared that il-4 contributed to their death and the il-4 mediated toxicity was confirmed in normal immunocompetent mice. the toxicity was re ...19921515552
induction of antigen-specific class i-restricted cytotoxic t cells by soluble proteins in vivo.cytotoxic t lymphocytes (ctl) are induced specifically against viral and tumor antigens presented by major histocompatibility complex class i molecules on the surface of infected or transformed cells. intracellular synthesized antigens are processed and associated with class i antigens within cells before presentation on the cell surface. because of this special requirement for ctl induction, exogenous soluble antigens do not, in general, induce specific ctl responses. to overcome this problem, ...19921518862
temperature-sensitive mutations in the vaccinia virus h4 gene encoding a component of the virion rna polymerase.four previously isolated temperature-sensitive (ts) mutants of vaccinia virus wr (ts1, ts31, ts55, and ts58) comprising a single complementation group (r. c. condit, a. motyczka, and g. spizz, virology 128:429-443, 1983) have been mapped by marker rescue to the h4l open reading frame located within the genomic hindiii-h dna fragment. the h4 gene is predicted to encode a 93.6-kda polypeptide expressed at late times during infection. nucleotide sequence alterations responsible for thermolabile gro ...19921527841
mutations in two cysteine-histidine-rich clusters in adenovirus type 2 dna polymerase affect dna binding.several point and linker insertion mutations in two cys-his-rich regions of adenovirus (ad) dna polymerase (pol) gene have been expressed in recombinant vaccinia virus. the resulting mutant enzymes were analyzed in vitro for their effects on dna synthesis activity, on ad-specific initiation assays, on gel shifts of ad origin sequences, and on interactions with adenovirus preterminal protein (ptp) and nuclear factor i (nfi). in general, mutants in downstream cys-his sequences had a pronounced eff ...19921527843
human immunodeficiency virus type 1 envelope glycoprotein molecules containing membrane fusion-impairing mutations in the v3 region efficiently undergo soluble cd4-stimulated gp120 release.the ability of soluble forms of cd4 to induce gp120 release from the human immunodeficiency virus type 1 envelope glycoprotein complex may reflect molecular events associated with membrane fusion. the third hypervariable (v3) region of gp120 appears to play a role in fusion independent of cd4 binding. we demonstrate herein that envelope glycoprotein molecules rendered fusion defective by mutations in conserved residues within the v3 region nevertheless undergo efficient soluble cd4-induced gp120 ...19921527858
proinsulin processing by the subtilisin-related proprotein convertases furin, pc2, and pc3.experiments using recombinant vaccinia viruses expressing rat proinsulin i coinfected into cos-7 cells with recombinant vaccinia virus expressing human furin, human pc2, mouse pc3 (subtilisin-related proprotein convertases 1-3, respectively), or yeast kex2 indicate that in this system both kex2 and furin produce mature insulin, whereas pc2 selectively cleaves proinsulin at the c-peptide-a-chain junction. this is a property consistent with its probable identity with the rat insulinoma granule typ ...19921528899
further analyses of the orthopoxviruses volepox virus and raccoon poxvirus.volepox virus (vpx) from skin lesions on a vole and a piñon mouse caught in california and raccoon poxvirus (rcn) from raccoons trapped in maryland were examined to begin elucidating their relationship to other orthopoxviruses, most of which are not known to be indigenous to the americas. vpx and rcn produced pinpoint, nonhemorrhagic pocks on chick embryo chorioallantoic membranes. in cell cultures both viruses produced 1-mm diameter, irregular plaques, a-type inclusions (atis), and despite prod ...19921529541
sequences of the raccoon poxvirus hemagglutinin protein.primers based on sequences flanking the vaccinia virus (vv) strain ihd hemagglutinin protein (ha) open reading frame (orf) enabled amplification of ha dna segments from the genome of raccoon poxvirus (rcn) and vv strain wr. the amplified segments produced unequal cross-hybridization signal intensities against each other, indicating sequence differences between the ha of rcn (in hindiii-g) and that of vv-wr (in hindiii-a). about 1.5 kb of sequences in the ha region were then determined from clone ...19921529542
introduction of foreign dna into the vaccinia virus genome by in vitro ligation: recombination-independent selectable cloning vectors.homologous recombination has been the exclusive means of introducing foreign dna into the genomes of large dna viruses. here we demonstrate that direct in vitro ligation can be used to efficiently insert dna fragments of up to 26,000 bp into the genome of vaccinia virus modified to contain a single noti site either in the escherichia coli lacz gene or in the vaccinia virus thymidine kinase gene. viruses containing chimeric genomes can be identified by chromogenic screening or thymidine-kinase-ne ...19921529553
protection of macaques against siv infection by subunit vaccines of siv envelope glycoprotein gp160.simian immunodeficiency virus (siv) is a primate lentivirus related to human immunodeficiency viruses and is an etiologic agent for acquired immunodeficiency syndrome (aids)-like diseases in macaques. to date, only inactivated whole virus vaccines have been shown to protect macaques against siv infection. protective immunity was elicited by recombinant subunit vaccines. four macaca fascicularis were immunized with recombinant vaccinia virus expressing sivmne gp160 and were boosted with gp160 pro ...19921531159
cleavage of the dengue virus polyprotein at the ns3/ns4a and ns4b/ns5 junctions is mediated by viral protease ns2b-ns3, whereas ns4a/ns4b may be processed by a cellular protease.the cleavage mechanism utilized for processing of the ns3-ns4a-ns4b-ns5 domain of the dengue virus polyprotein was studied by using the vaccinia virus expression system. recombinant vaccinia viruses vns2b-ns3-ns4a-ns4b-ns5, vns3-ns4a-ns4b-ns5, vns4a-ns4b-ns5, and vns4b-ns5 were constructed. these recombinants were used to infect cells, and the labeled lysates were analyzed by immunoprecipitation. recombinant vns2b-ns3-ns4a-ns4b-ns5 expressed the authentic ns3 and ns5 proteins, but the other reco ...19921531368
characterization of a secreted glycoprotein of the immunoglobulin superfamily inducible by mitogen and oncogene.the t1 gene is transiently activated by the ha-ras (ej) or v-mos oncoproteins and by mitogens in nih3t3 fibroblasts. its primary gene product of 337 amino acids (38 kda) undergoes extensive post-translational modification. for biochemical analysis, the t1 gene product was over-expressed in a vaccinia virus system. cells infected with a recombinant t1-vaccinia virus produce and secrete multiple proteins of 60-70 kda which react with polyclonal antisera raised against two t1-specific peptides. two ...19921532358
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 ...19921533596
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 ...19921534850
protection against friend retrovirus-induced leukemia by recombinant vaccinia viruses expressing the gag gene.high sequence variability in the envelope gene of human immunodeficiency virus has provoked interest in nonenvelope antigens as potential immunogens against retrovirus infection. however, the role of core protein antigens encoded by the gag gene in protective immunity against retroviruses is unclear. by using recombinant vaccinia viruses expressing the friend murine leukemia helper virus (f-mulv) gag gene, we could prime cd4+ t-helper cells and protectively immunize susceptible strains of mice a ...19921534853
immune response to the nominal phosphoprotein of rabies virus.the immune response to the nominal phosphoprotein (ns protein) of rabies virus was investigated with the use of a vaccinia recombinant virus that expressed the ns protein of a fixed rabies virus strain. mice of the h-2k haplotype that were injected with either live rabies virus or the vaccinia recombinant virus developed a strong cytolytic t-cell response specific for the ns protein. this response was under immune response (ir) gene control. the ns protein as presented by the vaccinia recombinan ...19921534957
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 ...19921536839
nuclear factor i enhances adenovirus dna replication by increasing the stability of a preinitiation complex.nuclear factor i (nfi) or its isolated dna-binding domain (nfi-bd) enhances initiation of adenovirus dna replication up to 50-fold at low concentrations of the precursor terminal protein-dna polymerase (ptp-pol) complex. both in solution and when bound to dna, nfi-bd can form a complex with ptp-pol. to investigate the mechanism of enhancement by nfi, we determined the stability of a functional preinitiation complex formed in vitro between ptp-pol and the origin. challenge experiments with a dist ...19921537346
the myxoma virus thymidine kinase gene: sequence and transcriptional mapping.the myxoma virus thymidine kinase (tk) gene is encoded on a 1.6 kb saci-sali restriction fragment located between 57.7 and 59.3 kb on the 163 kb genomic map. the nucleotide sequence of this fragment as well as 228 bp from the adjacent sali-aa2 fragment was determined and found to encode four major open reading frames (orfs). three of these orfs are similar in nucleotide sequence to orfs l5r and j1r, and the tk gene of vaccinia virus (vv). the fourth orf, mf8a, shows similarity to the orfs found ...19921538190
construction of vaccinia virus recombinants expressing several measles virus proteins and analysis of their efficacy in vaccination of mice.measles virus genes encoding the haemagglutinin (ha), fusion protein (f) or nucleoprotein (np) have been inserted into the vaccinia virus genome either alone or in various combinations. in each case the measles virus genes were expressed from the 7.5k promoter and were incorporated into the thymidine kinase (tk) or k1l loci of the copenhagen strain of vaccinia virus. cells infected by the recombinants synthesized measles virus proteins indistinguishable from those induced in measles virus-infect ...19921538193
production and characterization of recombinant mouse neurotrophin-3.neurotrophin-3 (nt-3) is a neurotrophic-factor that has recently been cloned on the basis of its structural similarity to other members of the nerve growth factor (ngf) gene family. in order to produce this protein, a recombinant vaccinia virus containing the coding region for mouse nt-3 was constructed. conditioned medium harvested from cells infected with the recombinant vaccinia virus contained biologically active nt-3 that was purified by chromatography on controlled-pore glass and reversed- ...19921541287
[construction of recombinant vaccinia virus for cdna-directed expression and its application]. 19921542737
a vaccinia serine protease inhibitor which prevents virus-induced cell fusion.a deletion mutant lacking the non-essential vaccinia virus gene k2l, a member of the serine protease inhibitor superfamily, was constructed. this virus replicates in vitro in all cell types tested and its virulence and immunogenicity in vivo are comparable to those of the parent virus in intranasally inoculated mice. however, in a variety of cell lines the cytopathic effect of the deletion mutant (vkl4) is markedly different from that caused by the parent virus: the absence of k2l in infected ce ...19921545218
a freeze-frame view of eukaryotic transcription during elongation and capping of nascent mrna.ribonuclease footprinting of nascent messenger rna within ternary complexes of vaccinia rna polymerase revealed an rna binding site that encompasses an 18-nucleotide rna segment. the dimensions of the binding site did not change as the polymerase moved along the template. capping of the 5' end of the rna was cotranscriptional and was confined to nascent chains 31 nucleotides or greater in length. purified capping enzyme formed a binary complex with rna polymerase in solution in the absence of nu ...19921546295
restricted replication of ectromelia virus in cell culture correlates with mutations in virus-encoded host range gene.ectromelia virus (strain moscow) was shown to replicate poorly or not at all in cell lines derived from the rabbit or hamster. the failure of ectromelia virus to replicate in cell lines derived from the hamster suggested that the virus lacked a functional cho host range (hr) gene required for multiplication in these cells. a dna fragment which hybridized to the cho hr gene was cloned from the ectromelia virus genome and shown by sequence analysis to be deleted of 506 bp within the ectromelia vir ...19921546448
in vivo recognition of orf virus early transcriptional promoters in a vaccinia virus recombinant.the 4.4-kb bamhi-e fragment of the orf virus (ov) genome contains three discrete open reading frames designated orf-pp, orf-1, and orf-3, all of which are flanked by vaccinia virus-like early transcriptional control sequences. to determine whether the vaccinia transcriptional machinery would recognize these promoters and faithfully transcribe ov genes in vivo the bamhi-e fragment was inserted into the thymidine kinase (tk) locus of vaccinia virus and the recombinant used in transcription studies ...19921546449
immature viral envelope formation is interrupted at the same stage by lac operator-mediated repression of the vaccinia virus d13l gene and by the drug rifampicin.specific missense mutations of the vaccinia virus d13l gene confer resistance to the effects of rifampicin on virion morphogenesis. we constructed a recombinant vaccinia virus in which elements of the escherichia coli lac operator system were used to regulate the d13l gene. replication of the recombinant vaccinia virus was dependent on addition of the inducer isopropyl beta-d-thiogalactoside (iptg) and the virus yield was decreased by more than 99% when iptg was omitted. under the nonpermissive ...19921546459
selective suppression of the catalytic activity of cdna-expressed cytochrome p4502b1 toward polycyclic hydrocarbons in the microsomal membrane: modification of this effect by specific amino acid substitutions.human hepatoma hepg2 cells were infected with recombinant vaccinia virus vectors containing cdnas encoding both known and variant rat cytochromes p450 (cyp). cyp2b1 and cyp2b2 cytochromes were equally well expressed (110-140 pmol/mg of microsomal protein) and catalyzed metabolism of 7,12-dimethylbenz[a]anthracene (dmba). their regioselectivity for dmba metabolism paralleled that of the respective purified rat liver enzymes and reproduced previously reported regioselective differences between cyp ...19921547225
sequence requirements for induction of cytolysis by the t cell antigen/fc receptor zeta chain.the zeta chain of the t cell antigen receptor is a dimeric transmembrane protein with a very short extracellular domain and an extended cytoplasmic tail that triggers t cell effector function when aggregated by extracellular stimuli. we have reduced the active site of zeta to an 18 residue motif that can be appended to the intracellular domain of other transmembrane proteins to endow them with receptor-like activity. the compact size of the motif appears to eliminate zeta mechanisms based on enz ...19921547489
identification of multiple srf n-terminal phosphorylation sites affecting dna binding properties.human serum response factor (srf) bearing a histidine tag was expressed using vaccinia virus. the recombinant protein was purified and shown to be phosphorylated mainly in its n-terminal part. the corresponding phosphorylation sites were mapped by microsequencing and also appear to be phosphorylated in endogenous serum response factor. four phosphorylation sites are located on serines within amino acids 77-85, while another phosphorylation site has been identified at ser103. mutations that consi ...19921547771
the hantaan virus m-segment glycoproteins g1 and g2 can be expressed independently.the two glycoproteins of hantaan virus (htv), g1 and g2, are encoded as a continuous single open reading frame in the m segment of the virion rna. they are believed to be synthesized contemporaneously via a polypeptide precursor which is then processed to yield two glycoproteins, both of which appear in the golgi complex of the cell. to study the properties of g1 and g2 as separate entities, we have constructed vaccinia virus recombinants which contain the sequences for each glycoprotein individ ...19921548747
activation of the human immunodeficiency virus type 1 long terminal repeat by vaccinia virus.a variety of dna viruses are known to activate gene expression directed by the long terminal repeat (ltr) of human immunodeficiency virus type 1 (hiv-1). in light of the proposed use of recombinant vaccinia virus for hiv-1 vaccines, evaluation of the role of vaccinia virus in hiv-1 activation is warranted. to investigate whether vaccinia virus induces hiv ltr-directed gene expression, transient expression assays in jurkat cells persistently infected with vaccinia virus (jvac) using plasmid dna c ...19921548751
an orthopoxvirus serpinlike gene controls the ability of infected cells to fuse.most orthopoxviruses encode a functional hemagglutinin (ha), which is nonessential for virus growth in cell culture. however, inactivation of the ha gene leads to the formation of polykaryocytes (syncytia) by fusion of infected cells at neutral ph. fusion is not observed when a functional ha gene is present. deletion of open reading frames (orfs) k2, k3, and k4 within the hindiii k fragment of the ha-positive (ha+) vaccinia virus strain wr also led to fusion of cells upon infection at neutral ph ...19921548753
vaccinia virus-specific human cd4+ cytotoxic t-lymphocyte clones.vaccinia virus-specific cytotoxic t-lymphocyte (ctl) clones were established from a healthy donor, who had been immunized with vaccinia virus vaccine, by stimulation of peripheral blood lymphocytes with uv-inactivated vaccinia virus antigen. the phenotype of all of the clones established was cd3+ cd4+ cd8- leu11-. we used a panel of allogenic vaccinia virus-infected b-lymphoblastoid cell lines and demonstrated that some of the clones recognized vaccinia virus epitopes presented by human leukocyt ...19921548761
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 ...19921548766
expansion and clonal deletion of peripheral t cells induced by bacterial superantigen is independent of the interleukin-2 pathway.injection of the bacterial superantigen staphylococcus aureus enterotoxin b (seb) into mice provokes a rapid expansion and subsequent contraction of the pool of seb-reactive t cells bearing t cell receptor (tcr) v beta 8 gene products. given that interleukin 2 (il-2) stimulates proliferation, abolishes anergy, and counteracts apoptotic cell death in t cells in vitro, we tested whether the il-2 synthesis inhibitor cyclosporin a (csa) or a vaccinia virus recombinant releasing high amounts of human ...19921551401
site-directed mutagenesis of a conserved domain in vaccinia virus thymidine kinase. evidence for a potential role in magnesium binding.alignment of prokaryotic and vertebrate type ii thymidine kinases (tk) (ec 2.7.1.21), such as that encoded by vaccinia virus (vvtk), reveals three conserved regions: designated domains i, iii, and vii. domains i and iii of vvtk contain residues which closely resemble segments a (atp) and b (mg2+), respectively, of a mg.atp binding descriptor proposed previously (walker, j.e., saraster, m., runswick, m-j., and gay, n.j. (1982) embo j. 1, 945-951). in support of this hypothesis, domain i of the vv ...19921551887
membrane interactions of synthetic peptides corresponding to amphipathic helical segments of the human immunodeficiency virus type-1 envelope glycoprotein.the human and simian immunodeficiency virus envelope glycoproteins, which mediate virus-induced cell fusion, contain two putative amphipathic helical segments with large helical hydrophobic moments near their carboxyl-terminal ends. in an attempt to elucidate the biological role of these amphipathic helical segments, we have synthesized peptides corresponding to residues 768-788 and 826-854 of hiv-1/wmj-22 gp160. circular dichroism studies of the peptides showed that the alpha helicity of the pe ...19921551918
site-directed mutagenesis of cytochrome p450s cyp2a1 and cyp2a2: influence of the distal helix on the kinetics of testosterone hydroxylation.cytochrome p450s cyp2a1 and cyp2a2 exhibit 88% sequence similarity, yet cyp2a1 metabolizes testosterone almost exclusively (90%) at the 7 alpha-position, whereas cyp2a2 forms several metabolites, with 15 alpha-hydroxytestosterone as a major metabolite. one of the regions with relatively low sequence homology corresponds by sequence alignment to the i and j helices of p450cam. since this region is known to be part of the active site for p450cam, 26 single point and two double point mutants were p ...19921554718
stability of ternary transcription complexes of vaccinia virus rna polymerase at promoter-proximal positions.we have used dna templates containing a vaccinia early promoter fused to g-less cassettes of varying length to study the formation of ternary transcription complexes by vaccinia virus rna polymerase. elongating polymerases were induced to pause at discrete sites on the dna template by omission of gtp from transcription reactions. for most of the templates examined, the predominant sites of pausing were at or near the downstream border of the g-less transcription unit, as revealed by the size dis ...19921559999
sickness and recovery of dogs challenged with a street rabies virus after vaccination with a vaccinia virus recombinant expressing rabies virus n protein.dogs were vaccinated intradermally with vaccinia virus recombinants expressing the rabies virus glycoprotein (g protein) or nucleoprotein (n protein) or a combination of both proteins. the dogs vaccinated with either the g or g plus n proteins developed virus-neutralizing antibody titers, whereas those vaccinated with only the n protein did not. all dogs were then challenged with a lethal dose of a street rabies virus, which killed all control dogs. dogs vaccinated with the g or g plus n protein ...19921560518
molecular attenuation of vaccinia virus: mutant generation and animal characterization.these studies demonstrated that the inbred balb/c mouse strain can be optimized for the assessment of vaccinia virus virulence, growth, and spread from the site of inoculation and immune protection from a lethal vaccinia virus challenge. the studies established that manipulation of the vaccinia virus genome generated mutants exhibiting a wide range of attenuated phenotypes. the nine nycbh vaccinia virus mutants had intracranial 50% lethal doses that ranged from 2 to greater than 7 log10 units. t ...19921560521
the vaccinia virus b1r gene product is a serine/threonine protein kinase.the nucleotide sequence of the vaccinia virus open reading frame b1 predicts a polypeptide with significant sequence similarity to the catalytic domain of known protein kinases. to determine whether the b1r polypeptide is a protein kinase, we have expressed it in bacteria as a fusion with glutathione s-transferase. affinity-purified preparations of the fusion protein were found to undergo autophosphorylation and also phosphorylated the exogenous substrates casein and histone h1. mutation of lysi ...19921560522
regulated expression of foreign genes in vaccinia virus under the control of bacteriophage t7 rna polymerase and the escherichia coli lac repressor.the gene encoding bacteriophage t7 rna polymerase (t7gene1) was placed under the control of regulatory elements from the escherichia coli lac operon to construct an inducible vaccinia virus expression system consisting entirely of prokaryotic transcriptional machinery. regulated expression of t7 rna polymerase was necessary to construct a stable recombinant vaccinia virus harboring a t7 promoter; otherwise, uncontrolled expression led to interference with endogenous virus replication. to this en ...19921560532
characterization of a 7-kilodalton subunit of vaccinia virus dna-dependent rna polymerase with structural similarities to the smallest subunit of eukaryotic rna polymerase ii.a previously unrecognized 7-kda polypeptide copurified with the dna-dependent rna polymerase of vaccinia virus virions. internal amino acid sequences of the small protein matched a viral genomic open reading frame of 63 codons. antipeptide antiserum was used to confirm the specific and complete association of the 7-kda protein with rna polymerase. the amino acid sequence predicted from the viral gene, named rpo7, was 23% identical to that of the smallest subunit of saccharomyces cerevisiae rna p ...19921560534
active immunotherapy with viral lysates of micrometastases following surgical removal of high risk melanoma.patients who develop lymph node metastases from melanoma are known to be at risk of developing further recurrences from melanoma following surgical removal of lymph node metastases. the purpose of the present study was to examine whether immunization over a 2 year period with a vaccine made from vaccinia viral lysates of an allogeneic melanoma cell (vmcl), following surgical removal of lymph node metastases, would help prevent the development of distant metastases and improve survival from the d ...19921561807
protein synthesis in vaccinia virus-infected cells. i. effect of hypertonic shock recovery.human hep-2 cells were submitted to hypertonic shock (210 mm nacl) to block host protein synthesis before infection with vaccinia virus. with the start of infection, the medium isotonicity (116 mm nacl) was restored, and the effect of viral infection on the recovery of host polyribosomes and protein synthesis was studied. although host translation blockage was released together with infection, vaccinia virus did not affect immediately host protein synthesis. during the first hour of recovery, in ...19921562234
effect of neurotropin on the activation of the plasma kallikrein-kinin system.bradykinin (bk), an important mediator of allergic reactions and pain induction, is released by the activation of the plasma kallikrein-kinin (k-k) cascade. neurotropin is a biological material obtained from inflamed rabbit skin inoculated with vaccinia virus and is widely used clinically in japan as an effective agent for these disorders. since its mechanism of action is not clearly known, we have investigated the effects of neurotropin on the human plasma k-k system. in dextran sulfate-activat ...19921562287
vaccinia virus recombinants expressing chimeric proteins of human immunodeficiency virus and gamma interferon are attenuated for nude mice.we have developed a method for attenuating vaccinia virus recombinants by expressing a fusion protein of a lymphokine and an immunogen. chimeric genes were constructed that coded for gamma interferon (ifn-gamma) and structural proteins of the human immunodeficiency virus type 1 (hiv-1). in this study, we describe the biological and immunological properties of vaccinia virus recombinants expressing chimeric genes of murine or human ifn-gamma with glycoprotein gp120, gag, and a fragment of gp41. a ...19921565633
rna polymerase-associated transcription specificity factor encoded by vaccinia virus.vaccinia virus encodes a multisubunit dna-dependent rna polymerase (ec 2.7.7.6) that is packaged in the infectious virus particle. this polymerase was found to contain a submolar polypeptide of approximately 85 kda in addition to the core subunits, which consist of two larger and several smaller polypeptides. the polymerase containing the 85-kda polypeptide was separated from the core polymerase by column chromatography. although the core polymerase actively transcribed heterologous single-stran ...19921565650
nyvac: a highly attenuated strain of vaccinia virus.a highly attenuated vaccinia virus strain, nyvac (vp866), was derived from a plaque-cloned isolate of the copenhagen vaccine strain by the precise deletion of 18 open reading frames (orfs) from the viral genome. among the orfs deleted from nyvac (vp866) are two genes involved in nucleotide metabolism, the thymidine kinase (orf j2r) and the large subunit of the ribonucleotide reductase (orf i4l); the gene encoding the viral hemagglutinin (orf a56r); the remnant (orf a26l) of a highly expressed ge ...19921566575
identification of a temperature-sensitive mutant of vaccinia virus defective in late but not intermediate gene expression.the vaccinia virus conditional-lethal temperature-sensitive (ts) mutant tsc63 is defective in the synthesis of some but not all postreplicative proteins. synthesis of the temporal "intermediate" class of proteins was unaffected, whereas "late" proteins were absent at the nonpermissive temperature. at the dna level, dna synthesis was unaffected, but telomere resolution was severely inhibited. in order to identify the defective gene responsible for this ts defect, we performed marker rescue and dn ...19921566576
genetic manipulation of african swine fever virus: construction of recombinant viruses expressing the beta-galactosidase gene.homologous recombination is shown to be specifically induced in vero cells by infection with african swine fever (asf) virus. the frequency of recombination induced by asf virus infection between cotransfecting plasmids is comparable to that found after infection with the prototype poxvirus, vaccinia virus. the induction of recombination is accompanied by replication of the plasmid templates in the asf virus-infected cells. an asf virus insertion/expression plasmid vector containing the escheric ...19921566585
molecular cloning and expression of a transformation-sensitive human protein containing the tpr motif and sharing identity to the stress-inducible yeast protein sti1.a transformation-sensitive human protein (ief ssp 3521) that is 2-fold up-regulated in sv40-transformed mrc-5 fibroblasts has been purified by two-dimensional gel electrophoresis, microsequenced, and cdna cloned using oligodeoxyribonucleotides. the 2.1-kilobase cdna encodes a 543-amino acid protein with a calculated molecular mass of 62.6 kda and a calculated pi of 6.77. expression of the cdna in ama cells using the vaccinia virus expression system followed by two-dimensional gel electrophoresis ...19921569099
recombinant mouse zp3 inhibits sperm binding and induces the acrosome reaction.mammalian fertilization involves interactions of sperm surface receptors with ligands of the zona pellucida, an extracellular matrix surrounding the ovulated egg. in mouse, the zona is composed of three glycoproteins. one of them, zp3, participates in primary sperm binding and in the subsequent triggering of the sperm's acrosome reaction. considerable evidence suggests that carbohydrate determinants of zp3 are responsible for binding to sperm and may be important for acrosomal exocytosis. a full ...19921577197
a single amino acid substitution abolishes feedback inhibition of vaccinia virus thymidine kinase.exploitation of differences in the substrate specificity of the type i and type ii thymidine kinases (ec 2.7.1.21, tk) expressed by the herpesviridae and poxviridae (and human cells), respectively, has lead to the development of effective antiherpetic drugs such as acyclovir and gancyclovir. analysis of type i tk protein sequences reveals a consensus sequence which corresponds to domain iv of type ii tk proteins such as that encoded by vaccinia virus (vv). the type i descriptor (xpho - + + xpho) ...19921577811
poxvirus expression vectors. 19921582244
steroid hormone synthesis by a vaccinia enzyme: a new type of virus virulence factor.vaccinia virus open reading frame (orf) salf7l has 31% amino acid identity to human 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase (3 beta-hsd). here we show that salf7l encodes an active 3 beta-hsd, by the conversion of pregnenolone to the steroid hormone progesterone. the gene is transcribed early during infection into a 1.4 kb mrna from an initiation site 12 bp upstream of the orf. an antiserum raised against bacterially expressed salf7l immunoprecipitated a 38 kda polypeptide ...19921582424
cytoplasmic domain truncation enhances fusion activity by the exterior glycoprotein complex of human immunodeficiency virus type 2 in selected cell types.to investigate the glycoprotein determinants of viral cytopathology, we constructed chimeric env genes between a noncytopathic strain of human immunodeficiency virus type 2 (hiv-2), designated hiv-2/st, and a highly fusogenic and cytopathic variant derived from this virus. expression of the resulting chimeric glycoproteins indicated that efficient syncytium formation in the human t-cell line sup t1 mapped to the c-terminal region of the transmembrane (tm) glycoprotein subunit. in this region, th ...19921583738
control of rabies in wildlife. 19921585150
rna packaging signal of human immunodeficiency virus type 1.cells infected with a recombinant vaccinia virus carrying the gag and pol regions of the human immunodeficiency virus type 1 genome (vac-gag/pol) released human immunodeficiency virus (hiv)-like particles containing hiv-specific rna. however, cells infected with another recombinant vaccinia, vac-gag/pol-dp, derived through the deletion of an 85-base region (nucleotide positions 679-763) of the hiv genome between the primer binding site and the gag initiation codon of vac-gag/pol, produced hiv-li ...19921585635
mice immunized with a subviral particle containing the japanese encephalitis virus prm/m and e proteins are protected from lethal jev infection.extracellular subviral particles produced by hela cells infected with a recombinant vaccinia virus encoding the prm and e genes of japanese encephalitis virus (jev) were purified and characterized. these particles contained the jev prm/m and e proteins embedded in a lipid bilayer, and rna was not detected in particles using the polymerase chain reaction and primers recognizing a part of the jev e gene. the particles were uniformly spherical with a 20-nm diameter and had 5-nm projections on their ...19921585642
a constitutively expressed vaccinia gene encodes a 42-kda glycoprotein related to complement control factors that forms part of the extracellular virus envelope.nucleotide sequence analysis of a 42-kb region of the vaccinia virus (strain western reserve) genome identified a gene with the potential to encode a 35.1-kda polypeptide with properties of a membrane glycoprotein (smith et al., j. gen. virol. 72, 1349-1376, 1991). the 317 amino acid open reading frame (orf) has similarity with complement control proteins and a secretory vaccinia virus protein (c28k) which interferes with complement function. the predicted b5r gene product differs from the latte ...19921585649
vaccinia virus-mediated inhibition of host protein synthesis involves neither degradation nor underphosphorylation of components of the cap-binding eukaryotic translation initiation factor complex eif-4f.recent reports indicated that vaccinia virus late mrnas contain a unique 5' poly(a) leader sequence and that the in vitro translation of these mrnas may be relatively cap-independent. these observations led us to examine the possibility that the mechanism of inhibition of host protein synthesis by vaccinia virus, like that of certain other viruses, involves specific modifications of the cap-binding translation initiation factor complex eif-4f. the eif-4f complex was affinity-purified from human ...19921585660
the eukaryotic translation initiation factor 4e is not modified during the course of vaccinia virus replication.the ability of vaccinia virus to inhibit processes of cap-dependent translational initiation by inactivating the eukaryotic translation initiation factor 4e (eif-4e) has been examined. analyses of the quantities of eif-4e present in either uninfected mouse l929 cells or vaccinia virus-infected cells showed that during the first 12 hr of virus replication, when there is a marked decrease in host gene expression in infected cells, there is no change in the total amount of eif-4e present. analyses ...19921585661
inability of il-2 and il-10 to counteract b cell clonal deletion.the b cell antigen receptor (bcr) delivers inhibitory signals in nascent b cells leading to the establishment of tolerance via clonal deletion or clonal anergy depending upon the type of antigen to which the b cells are exposed. in previous work, it has been demonstrated that activated th2 cells, as well as some recombinant lymphokines, prevent the inhibition of growth and subsequent cell death induced through the bcr in model b cell lymphomas. herein, we extend this work to another th2 lymphoki ...19921586962
vaccinia recombinants as vaccine vectors. 19921587539
vaccinia virus gene salf5r is non-essential for virus replication in vitro and in vivo.a gene designated salf5r from within the sali f fragment of vaccinia virus strain wr has been characterized. the predicted primary translation product has 194 amino acids with an mr of 22578 and has features typical of a class i membrane glycoprotein. at the amino and carboxy termini there are runs of hydrophobic residues that might function as membrane signal and anchor sequences, respectively, and after the c-terminal hydrophobic sequence there is a short charged sequence that may prevent pass ...19921588324
detection of rna-protein complex in vaccinia virus core in vitro transcription system.the incubation of vaccinia virus cores in appropriate conditions promotes the release of core proteins into a supernatant fraction. under transcription assay conditions core mrnas are extruded in association with viral core proteins, however the presence of these proteins within the core particle is not essential for rna synthesis and extrusion. the rna-protein complex is resistant to micrococcal nuclease. five proteins of 60k, 43k, 28k, 18k and 14.5k with rna-binding abilities have been identif ...19921588325
a 2-year follow-up of an anti-hiv immune reaction in hiv-1 gp160-immunized healthy seronegative humans: evidence for persistent cell-mediated immunity.the first trial of an anti-hiv immunization, using a recombinant vaccinia virus expressing gp160 (rv) for priming and paraformaldehyde-fixed rv-infected pbls and soluble gp 160 for boosting, clearly showed an in vitro hiv-protective immune reaction. this result led us to carry out an additional 2 year phase i clinical trial in 25 hiv-seronegative volunteers, using hiv gp 160 antigens for immunization in four different protocols. the 2 year trial showed (a) the safety of the preparations, (b) a t ...19921588488
pulmonary eosinophilic response to respiratory syncytial virus infection in mice sensitized to the major surface glycoprotein g.to investigate the contribution of immunity to individual respiratory syncytial (rs) virus proteins to the augmentation of pulmonary pathology, mice were scarified with recombinant vaccinia viruses (rvv) expressing individual rs virus proteins. the pulmonary response to infection with rs virus was monitored by bronchoalveolar lavage (bal). in mice vaccinated with the major surface glycoprotein (g), 14-25% of bal cells were eosinophils; these comprised less than 3% of bal cells from other groups ...19921591217
changes in muscarinic acetylcholine receptors in the isolated duodenum from repeatedly cold-stressed rats and the effect of neurotropin.in rats repeatedly cold-stressed by specific alternation of rhythm in environmental temperature (sart-stressed rats), the contractile response to acetylcholine (ach) of the isolated duodenum was remarkably decreased, whereas the contractile responses to k+, ba2+ and ca2+ were comparable to those in non-stressed rats. the amount of [3h]quinuclidinyl benzilate in the duodenum of sart-stressed rats was about 50% of that in non-stressed rats, but the kd value remained unchanged. long-term administra ...19921593927
proprotein conversion is determined by a multiplicity of factors including convertase processing, substrate specificity, and intracellular environment. cell type-specific processing of human prorenin by the convertase pc1.proprotein and prohormone processing at pairs of basic residues is generally thought to be both tissue- and precursor-specific and to be developmentally regulated. furin, pc1 (also called pc3), and pc2 represent three recently discovered subtilisin-like proteinases which cleave a number of precursors at the same pairs of basic residues normally processed in vivo. using human prorenin as a model, we show that pc1 can process it to active renin in cells containing secretory granules, such as the s ...19921597471
primate responses to a vaccinia-rabies glycoprotein recombinant virus vaccine.over the past decade, a vaccinia-rabies glycoprotein (v-rg) recombinant vaccine virus has been developed for the potential control of wildlife rabies through oral immunization via baits. prior to widespread field applications in varying ecological settings, extensive laboratory safety testing is necessary in a number of target and non-target species to quantify real or perceived risks and to monitor potential adverse health effects. moreover, in order to mitigate concerns over inadvertent human ...19921598786
monoclonal antibodies to rat liver microsomal cytochrome b5.hybridomas obtained by the fusion of spleen cells from rat cytochrome b5-immunized mice with mouse myeloma cells produced five groups of monoclonal antibodies (mabs) with three mouse immunoglobulin subtypes: igg1, igg2b and igm. all of the mabs bound strongly to rat cytochrome b5 as measured by radioimmunoassay (ria). four clones of mabs were also strongly immunoreactive with cytochrome b5 when tested by western blotting, but only one of the mabs (1-39-2) weakly immunoprecipitated cytochrome b5 ...19921599506
vaccinia virus vectors. 19921600384
the transmembrane glycoprotein of human immunodeficiency virus type 1 induces syncytium formation in the absence of the receptor binding glycoprotein.to study the intracellular transport and biological properties of the human immunodeficiency virus type 1 (hiv-1) transmembrane glycoprotein (tm; gp41), we constructed a truncated envelope gene in which the majority of the coding sequences for the surface glycoprotein (su; gp120) were deleted. transient expression of this truncated env gene in primate cells resulted in the biosynthesis of two proteins with m(r)s of 52,000 and 41,000, respectively. immunofluorescence studies with antibodies to th ...19921602536
overexpression of the influenza virus polymerase can titrate out inhibition by the murine mx1 protein.the murine mx1 protein is an interferon-inducible protein which confers selective resistance to influenza virus infection both in vitro and in vivo. the precise mechanism by which the murine mx1 specifically inhibits replication of influenza virus is not known. previously, sensitive replication systems for influenza virus ribonucleoprotein, in which a synthetic influenza virus-like ribonucleoprotein is replicated and transcribed by influenza virus proteins provided in trans, have been developed. ...19921602538
role of cell-associated enveloped vaccinia virus in cell-to-cell spread.the roles of intracellular naked (inv), cell-associated enveloped (cev), and extracellular enveloped (eev) forms of vaccinia virus in cell-to-cell and longer-range spread were investigated by using two closely related strains of vaccinia virus, wr and ihd-j. we confirmed previous results that wr and ihd-j produced similar amounts of inv and formed similar-size primary plaques but that ihd-j produced 10 to 40 times more eev and spread to distant cells much more efficiently than did wr. neverthele ...19921602540
vaccinia virus b1 kinase: phenotypic analysis of temperature-sensitive mutants and enzymatic characterization of recombinant proteins.the vaccinia virus b1 gene encodes a 34-kda protein with homology to protein kinases. in l cells infected nonpermissively with mutants containing lesions in the b1 gene (ts2 and ts25), the infectious cycle arrests prior to dna replication. in this report, we demonstrate that dna synthesis ceases when cultures infected with these mutants at 32 degrees c are shifted to the nonpermissive temperature (39.5 degrees c) in the midst of dna replication. we also show that b1 protein is synthesized transi ...19921602551
pollen-derived rice calli that have large deletions in plastid dna do not require protein synthesis in plastids for growth.albino rice plants derived from pollen contain plastid genomes that have suffered large-scale deletions. from the roots of albino plants, we obtained several calli containing homogeneous plastid dna differing in the size and position of the deletion. dna differing in the size and position of the deletion. southern blotting and pulsed field gel electrophoresis experiments revealed that the dnas were linear molecules having a hairpin structure at both termini, existing as monomers (19 kb) or dimer ...19921603057
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