| apoptosis reduces both the in vitro replication and the in vivo infectivity of a baculovirus. | apoptotic programmed cell death occurs when the insect cell line sf-21, derived from spodoptera frugiperda, is infected with mutants of the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) which lack a functional p35 gene. however, infection of the trichoplusia ni tn-368 cell line with p35 mutants does not result in apoptosis (r. clem, m. fechheimer, and l. miller, science 254:1388-1390, 1991). we have examined the effect of apoptosis on acmnpv infections in cell lines and ... | 1993 | 8510202 |
| in vitro transcription from baculovirus late gene promoters: accurate mrna initiation by nuclear extracts prepared from infected spodoptera frugiperda cells. | extracts prepared from nuclei of autographa californica nuclear polyhedrosis virus-infected spodoptera frugiperda cells were shown to support in vitro transcription from baculovirus late gene promoters. in vitro transcription was optimized for the late promoter of the 39k gene. the mg2+ concentration was critical; concentrations higher than 1 to 2 mm did not support late transcription. additional conditions included template (40 micrograms/ml), extract (2.5 mg/ml), and incubation time (25 min). ... | 1993 | 8510206 |
| a baculovirus-expressed fusion protein containing the antibody-binding domain of protein a and insect luciferase. | a fusion construct encoding two antibody-binding sites of protein a from staphylococcus aureus and click beetle, pyrophorus plagiophthalamus, luciferase (lucgr) was designed and expressed using the baculovirus system. the construct was inserted under the transcriptional regulation of the polyhedrin gene promoter of the autographa californica nuclear polyhedrosis virus (acnpv) and expressed in the insect spodoperta frugiperda cell line during viral infection. the properties of the resultant chime ... | 1993 | 8512706 |
| a method for producing recombinant baculovirus expression vectors at high frequency. | a system has been developed that can generate recombinant baculovirus expression vectors at frequencies approaching 100%. this system provides a selection for recombinant viruses by using the essential gene downstream of the autographa californica nuclear polyhedrosis virus (acmnpv) polyhedrin expression locus. two acmnpv derivatives were constructed in which the expression locus and part of the downstream gene are flanked by restriction sites. the parental viruses are viable; however, restricti ... | 1993 | 8512707 |
| a simple and efficient procedure for the oral inoculation of trichoplusia ni larvae with polyhedrin-negative recombinant baculovirus. | current procedures for inoculating lepidopteran larvae with polyhedrin-negative recombinant baculovirus, i.e. intracoelomic injection or coinfection with wild type virus, are laborious and can compromise final yields of recombinant protein. herein is described a simple and efficient method for oral inoculation. up to 100% infection was obtained when individual early fifth instar trichoplusia ni larvae were fed a small piece of a formaldehyde-free insect diet to which 4.2 x 10(5) pfu of a polyhed ... | 1993 | 8514841 |
| location of two putative origins of dna replication of autographa californica nuclear polyhedrosis virus. | previously, we described a defective autographa californica nuclear polyhedrosis virus (acmnpv) which must contain cis-acting elements required for dna synthesis, such as the origin(s) of replication (ori). defective genomes of acmnpv generated after serial undiluted passage were analyzed further. three small separated regions were retained in dna of defective acmnpv and accumulated in extracellular defective interfering viruses as well as in intracellular dna after 40 passages. two of these reg ... | 1993 | 8517035 |
| expression and radiolabeling of human c-reactive protein in baculovirus-infected cell lines and trichoplusia ni larvae. | human c-reactive protein (crp) is a member of the pentraxin family of proteins which are molecules composed of five identical subunits arranged in a planar configuration. in the present study a human crp cdna clone was ligated into the baculovirus vector pvl1393 which was used to establish a recombinant strain of baculogold autographa californica multiple nuclear polyhedrosis virus containing the coding and leader sequence for human crp (designated acmnpv-crp). synthesis and secretion of crp wer ... | 1995 | 8527929 |
| construction and insect larval expression of recombinant vesicular stomatitis nucleocapsid protein and its use in competitive elisa. | the gene encoding the nucleocapsid (n) protein of indiana 1 serotype vesicular stomatitis virus (vsv-in1) was transferred into the genome of autographa californica nuclear polyhedrosis virus (baculovirus) as a full-length non-fusion construct under the control of the polyhedrin gene promoter. recombinant n protein was obtained from trichoplusia ni insect larvae inoculated 72-96 h previously with the recombinant baculovirus. polyclonal antibody (pab) against vsv-in1 was produced in mice using vsv ... | 1995 | 8530566 |
| convenient desktop-scale production of the extracellular domain of the human growth hormone receptor by an insect-baculovirus secretion system using a protein-free culture. | expression of a gene encoding the extracellular domain of the human growth hormone receptor (hghr-ed) inserted into the genome of autographa californica nuclear polyhedrosis virus was done using a desktop-scale spinner culture. spodoptera frugiperda 9 (sf9) cells infected with the recombinant virus secreted a protein with hgh-binding activity into the medium. oxygen supplementation was required for high level secretion of the product. the highest cell production capability was estimated at more ... | 1995 | 8534995 |
| identification and characterization of the v-cath gene of the baculovirus, cfmnpv. | the v-cath gene of the autographa californica multi-nucleocapsid nuclear polyhedrosis virus (acmnpv) encodes a cathepsin l-like proteinase which plays a role in the liquefaction of host tissues during a viral infection [1]. we have identified a homologous gene in the spruce budworm virus, choristoneura fumiferana mnpv (cfmnpv). the cfmnpv v-cath gene is 74% identical to acmnpv v-cath at the nucleotide sequence level and 80% identical at the level of predicted amino acid sequence. transcription a ... | 1995 | 8547313 |
| human cathepsin o2, a matrix protein-degrading cysteine protease expressed in osteoclasts. functional expression of human cathepsin o2 in spodoptera frugiperda and characterization of the enzyme. | cathepsin o2, a human cysteine protease predominantly present in osteoclasts, has been functionally expressed in spodoptera frugiperda sf9 cells using the autographa californica nuclear polyhedrosis virus. following in vitro activation at ph 4.0 with pepsin, active enzyme with an apparent molecular weight of 29,000 was obtained. n-terminal sequencing revealed the typical processing site for cysteine proteases of the papain family with a proline in the position adjacent to the n-terminal alanine ... | 1996 | 8567669 |
| production of polyhedra of the autographa californica nuclear polyhedrosis virus using the sf21 and tn5b1-4 cell lines and comparison with host-derived polyhedra by bioassay. | both wild-type and recombinant baculoviruses are becoming more attractive for the control of insect pests. thus, there is an increased incentive to address and resolve logistical problems associated with large-scale production of these viruses. in this study, we have compared the potential of two insect cell lines, tn5b1-4 and sf21, for the production of polyhedra and compared the efficacy of both cell culture-derived and host-derived viruses by bioassay. the efficacy of both wild-type acmnpv an ... | 1995 | 8568278 |
| nucleotide sequence of a 1446 base pair sali fragment and structure of a novel early gene of leucania separata nuclear polyhedrosis virus. | a 1446 bp sali fragment of lsnpv was sequenced by the silver staining method, and two large open reading frames (orfs, orf1 and orf2) were found, both contain typical characteristics of the 5' regulatory elements of baculovirus early genes. orf1 is 345 bp long with the capacity to encode a putative protein of 114 amino acid residues with mw about 13 kda and was designated p13 gene, orf2 comprises 248 bp from the 3' end of the fragment. in the untranslated region (utr) of orf1, a 33 bp mini cistr ... | 1995 | 8572949 |
| nucleocapsid protein n of lelystad virus: expression by recombinant baculovirus, immunological properties, and suitability for detection of serum antibodies. | the orf7 gene, encoding the nucleocapsid protein n of lelystad virus (lv), was inserted downstream of the p10 promoter into autographa californica nuclear polyhedrosis virus (baculovirus). the resulting recombinant baculovirus, designated bac-orf7, expressed a 15-kda protein in insect cells. this protein was similar in size to the n protein expressed by lv in cl2621 cells when it was analyzed on sodium dodecyl sulfate-polyacrylamide gels. the n protein expressed by bac-orf7 was immunoprecipitate ... | 1995 | 8574824 |
| two human alpha 2-adrenoceptor subtypes alpha 2a-c10 and alpha 2b-c2 expressed in sf9 cells couple to transduction pathway resulting in opposite effects on camp production. | the baculovirus expression vector system utilizing the strong polyhedrin gene promoter of the autographa californica nuclear polyhedrosis virus (acnpv) was used for high level expression of the two alpha 2-adrenoceptor subtypes alpha 2a-c10 and alpha 2b-c2 in spodoptera frugiperda (sf-9) insect cells. for rapid screening of recombinant viruses the luciferase gene was expressed under the early etl-promoter (early transcript large) in the same plasmid. both receptor subtypes showed the same rank o ... | 1995 | 8575536 |
| the baculovirus cysteine protease has a cathepsin b-like s2-subsite specificity. | autographa californica nuclear polyhedrosis virus (acnpv) encodes a functional cysteine protease of the papain family which is expressed after infection in spodoptera fruglperda sf9 cells. the protease displays an inhibition profile typical for cysteine proteases and is highly active against synthetic peptide substrates. the ph optimum of the bell-shaped ph-activity curve is between 5.0 and 5.5. the best substrate tested is z-arg-arg-mca which is specific for cathepsin b. the specificity constan ... | 1995 | 8590630 |
| an autographa californica nucleopolyhedrovirus lef-2 mutant: consequences for dna replication and very late gene expression. | in order to define factors involved in very late autographa californica nucleopolyhedrovirus (acmnpv) gene function, random mutagenesis of a baculovirus recombinant (acuw1.lacz) by 5'-bromodeoxyuridine treatment was performed. five viruses were selected with deficiencies in very late gene expression. these were characterized by complementation analysis. one mutant virus, vld1, was found to be completely deficient in very late gene function. this virus could be complemented by a helper virus to e ... | 1996 | 8599220 |
| identification and analysis of an autographa californica nuclear polyhedrosis virus structural protein of the occlusion-derived virus envelope: odv-e56. | an autographa californica nuclear polyhedrosis virus gene encoding an occlusion-derived virus (odv) envelope protein of 56 kda was identified and sequenced. transcription initiates from a conserved baculovirus late motif (ataag) with transcripts detected from 16 through 72 hr p.i. the protein is detected in infected cell extracts from 36 hr p.i. western blot assay of odv, bv, viral envelope, and nucleocapsid preparations coupled with immunoelectron microscopy reveal that this protein localizes t ... | 1996 | 8599240 |
| actin binding and proteolysis by the baculovirus acmnpv: the role of virion-associated v-cath. | infection of larvae by autographa californica m nuclear polyhedrosis virus (acmnpv) results in liquefaction of susceptible hosts, presumably due to the breakdown of cells and extracellular matrices. in spodoptera frugiperda tissue culture cells, infection leads to dramatic rearrangement and eventual destruction of the actin cytoskeleton. the first of these rearrangements is the formation of actin cables in the cytoplasm of the cell. cable formation requires release of the budded virus (bv) nucle ... | 1996 | 8607267 |
| baculovirus infection of spodoptera exigua larvae: lacz expression driven by promoters of early genes pe38 and me53 in larval tissue. | to follow the progression of infection of autographa californica multicapsid nuclear polyhedrosis virus (acmnpv) within tissues of its larval host, we have constructed acmnpv recombinants carrying lacz reporter genes under the control of the early virus promoters pe38 and me53, in addition to the authentic genes. the early promoter-lacz gene cassettes were located upstream of the very late polyhedrin gene. in infected insect cell lines, pe38 transcription is initiated at an early promoter, while ... | 1996 | 8609477 |
| the dna polymerase and helicase genes of a baculovirus of orgyia pseudosugata. | regions of the genome of the orgyia pseudotsugata multinucleocapsid nuclear polyhedrosis virus (opmnpv) containing the dna polymerase and helicase genes were sequenced. the dna polymerase and helicase genes encode predicted proteins of 985 (112.6 kda) and 1223 (140.5 kda) amino acids and exhibited 63 % and 59 % amino acid identity, respectively, with their homologues in the autographa californica mnpv (acmnpv). the influence of sequence variation between the opmnpv and acmnpv dna polymerase and ... | 1996 | 8609478 |
| studies of choristoneura fumiferana nuclear polyhedrosis virus gene expression in insect cells. | to investigate the mechanisms regulating baculovirus virulence and host range we have begun to study choristoneura fumiferana nuclear polyhedrosis virus (cfmnpv) and its gene expression in permissive and nonpermissive cells. we have identified and mapped three genes on the cfmnpv genome. the polyhedrin gene is located from 0.0 to 2.0 m.u. and two other genes, dnapol and p143, both of which are essential for baculovirus dna replication, are located from 35.3 to 40.9 m.u. and 55.5 to 63.4 m.u., re ... | 1996 | 8610448 |
| purification of a virus-induced rna polymerase from autographa californica nuclear polyhedrosis virus-infected spodoptera frugiperda cells that accurately initiates late and very late transcription in vitro. | the virus-induced rna polymerase from autographa californica nuclear polyhedrosis virus-infected spodoptera frugiperda cells was separated from the three host nuclear rna polymerases by deae-sephadex chromatography and then purified through two more steps: heparin-agarose chromatography and glycerol gradient ultracentrifugation. fractions from each of these purification steps have been assayed in vitro for the ability to perform accurate initiation of transcription on a late (p6.9) and a very la ... | 1996 | 8614978 |
| trans-activation of a cell housekeeping gene promoter by the ie1 gene product of baculoviruses. | protein ie1 is the product of a baculovirus gene, ie1, that is activated immediately upon entrance of the viral genome into the cell nucleus. this protein was previously shown to be a trans-regulator of viral genes whose products are required for initiation of the infectious cycle including viral dna replication. to test whether the ie1 protein is also capable of trans-regulating nuclear genes of the host in vitro and in vivo, we transfected the ie1 gene of bombyx mori nuclear polyhedrosis virus ... | 1996 | 8615012 |
| characterization of a highly conserved baculovirus structural protein that is specific for occlusion-derived virions. | a highly conserved baculovirus late gene called odvp-6e was shown to be a structural protein that is specific for occlusion-derived virus (odv) envelopes. the complete sequence of this gene is presented for both orgyia pseudotsugata nuclear polyhedrosis virus (opmnpv) and cydia pomonella granulosis virus (cpgv). the predicted sizes of the opmnpv and cpgv odvp-6e are 40, 241, and 38,655 respectively. the opmnpv odvp-6e gene was transcriptionally mapped and was shown to initiate from a consensus l ... | 1996 | 8615018 |
| the viral ubiquitin gene of autographa californica nuclear polyhedrosis virus is not essential for viral replication. | the baculovirus autographa californica nuclear polyhedrosis virus (acnpv) encodes a protein with significant homology to ubiquitin. to study the role of viral ubiquitin in infection, a recombinant virus was constructed with a frameshift mutation within the coding sequence of the viral ubiquitin gene, v-ubi. this recombinant, named vubi-fs, was viable, indicating that viral ubiquitin is not essential for replication in tissue culture. however, the yields of infectious budded virus were decreased ... | 1996 | 8615029 |
| functional expression of the aequorea victoria green fluorescent protein in insect cells using the baculovirus expression system. | a dna fragment encoding the green fluorescent protein (gfp) was isolated via pcr from a jellyfish aequorea victoria cdna, cloned and sequenced. subsequently, a recombinant baculovirus bearing the coding region of the gfp under the transcriptional control of the autographa californica nuclear polyhedrosis virus (acmnpv) polyhedrin gene promoter was constructed and isolated. high-level expression of gfp could be easily monitored in spodoptera frugiperda (sf9) insect cells after infection with reco ... | 1996 | 8619796 |
| physical map of anagrapha falcifera multinucleocapsid nuclear polyhedrosis virus. | a physical map of anagrapha falcifera multinucleocapsid nuclear polyhedrosis virus (afmnpv) dna was constructed for restriction endonucleases ecori, hindiii, psti and xhoi. the genome size was estimated to be 130 kbp. ordering of the restriction fragments was accomplished by cross-blot hybridization, double digestion and dna-dna hybridization. the polyhedrin gene and homologous repeat (hr) regions were located by hybridization to the autographa californica mnpv (acmnpv) polyhedrin gene and hr4, ... | 1996 | 8627218 |
| baculovirus-mediated gene transfer into mammalian cells. | this paper describes the use of the baculovirus autographa californica multiple nuclear polyhedrosis virus (acmnpv) as a vector for gene delivery into mammalian cells. a modified acmnpv virus was prepared that carried the escherichia coli lacz reporter gene under control of the rous sarcoma virus promoter and mammalian rna processing signals. this modified baculovirus was then used to infect a variety of mammalian cell lines. after infection of the human liver cell lines hepg2, >25% of the cells ... | 1996 | 8637876 |
| human bleomycin hydrolase: molecular cloning, sequencing, functional expression, and enzymatic characterization. | we have cloned the cdna of human bleomycin hydrolase (hbh), a protease which is thought to be involved in the metabolic inactivation of the antineoplastic drug bleomycin. the open reading frame consists of 1365 base pairs and is predicted to encode a 52 kda protein. the protein shares 40% identity with yeast bleomycin hydrolase and contains the conserved active site residues (cys, his, asn) characteristic for cysteine proteases of the papain superfamily. human bleomycin hydrolase has been functi ... | 1996 | 8639621 |
| cloning and biologic activities of a bovine interferon-alpha isolated from the epithelium of a rotavirus-infected calf. | a cdna encoding a distinct bovine (bo) interferon (ifn) alpha, designated boifn-alpha e, was generated from gut epithelial cells isolated from a rotavirus-infected calf. the boifn-alpha e cdna sequence shared a greater than 90% identity with the other boifn-alpha subtypes. the cdna encoding boifn-alpha e has been expressed in insect cells using the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) as a vector. insect cells infected with recombinant virus secreted a protein w ... | 1996 | 8640447 |
| identification of baculovirus gene that promotes autographa californica nuclear polyhedrosis virus replication in a nonpermissive insect cell line. | a gene that promotes autographa californica m nuclear polyhedrosis virus (acmnpv) replication in iplb-ld652y cells, a cell line that is nonpermissive for acmnpv, was identified in lymantria dispar m nuclear polyhedrosis virus (ldmnpv). cotransfection of acmnpv dna and a plasmid carrying the ldmnpv gene into iplb-ld652y cells results in acmnpv replication. the gene maps between 43.3 and 43.8 map units on the 162-kbp genome of ldmnpv. it comprises a 218-codon open reading frame and encodes a polyp ... | 1996 | 8642646 |
| factors regulating baculovirus late and very late gene expression in transient-expression assays. | eighteen genes of autographa californica nuclear polyhedrosis virus are necessary and sufficient to transactivate expression from the late vp39 promoter in transient-expression assays in sf-21 cells. these 18 genes, known as late expression factor genes (lefs), are also required to transactivate the very late promoter of the polyhedrin gene, polh, but expression from this promoter is relatively weak compared with expression from the vp39 promoter. to further define the factors required for late ... | 1996 | 8642657 |
| a new set of versatile vectors for the heterologous expression of foreign genes using the baculovirus system. | a new set of versatile advanced baculovirus (bv) vectors for the production of fused proteins in insect cells, under the control of the strong polyhedrin promoter of the autographa california nuclear polyhedrosis virus (acmnpv), has been constructed. the vectors contain peptide tags which allow immunological detection, as well as purification of recombinant protein produced via the bv expression system. | 1996 | 8647444 |
| baculovirus-directed expression of human prostatic steroid 5 alpha-reductase 1 in an active form. | in the human prostate, the enzyme steroid 5 alpha-reductase (h5 alpha r) catalyses the conversion of testosterone into the more potent androgen, dihydrotestosterone. two distinct cdnas coding for h 5 alpha r in the human prostate have been previously characterized. enzyme h5 alpha r1 shows a maximum activity at basic ph whereas h5 alpha r2 has an acidic ph optimum activity. we report here the expression of the human steroid h5 alpha r1 in a eukaryotic expression system: the baculovirus-directed- ... | 1993 | 8664165 |
| a recombination-efficient baculovirus vector for simultaneous expression of multiple genes. | the baculovirus system is an extremely powerful tool for expression of heterologous genes in eukaryotic environment. a multiple expression vector, pbacucmp3, was constructed which harbored two copies of the autographa californica nuclear polyhedrosis virus very late gene promoter and the drosophila melanogaster 70-kda heat-shock protein (hsp70) promoter with downstream unique restriction sites for cloning of three independent foreign genes. co-transfection of pbacucmp3 with bsu36i-linearized vir ... | 1996 | 8666274 |
| the gp64 envelope fusion protein is an essential baculovirus protein required for cell-to-cell transmission of infection. | to demonstrate the essential nature of the baculovirus gp64 envelope fusion protein (gp64 efp) and to further examine the role of this protein in infection, we inactivated the gp64 efp gene of autographa californica multicapsid nuclear polyhedrosis virus (acmnpv) and examined the biological properties of this virus in vivo. to provide gp64 efp during construction of the recombinant gp64 efp-null acmnpv baculovirus, we first generated a stably transfected insect cell line (sfpop64-6) that constit ... | 1996 | 8676487 |
| [sequencing of polyhedrin gene of leucania separata multiple capsid nuclear polyhedrosis virus by silver sequencing system]. | the genome of lsnpv was digested with ecorv and cloned into pbluescript. with the partial polyhedrin gene of autographa californica as a probe, positive recombinants were screened by colony in situ hybridization, further characterized by sequencing, and designated as plsev and plsph5. the intact 741 bp polyhedrin gene of lsnpv is located on the two recombinants revealed by silver sequencing system, and shares 80.0% and 97.0% nucleotide identity of the coding sequence, 87.8% and 97.5% amino acid ... | 1995 | 8701583 |
| abortive infection of the baculovirus autographa californica nuclear polyhedrosis virus in sf-9 cells after mutation of the putative dna helicase gene. | homologous recombination between the autographa californica nuclear polyhedrosis virus (acnpv) genome and a 0.6-kbp-long dna fragment derived from the putative dna helicase gene of bombyx mori nuclear polyhedrosis virus generates eh2-acnpv, an expanded-host-range acnpv mutant (s. maeda, s.g. kamita, and a. kondo, j. virol. 67:6234-6238, 1993). after inoculation at a high multiplicity of infection (moi), eh2-acnpv replicates efficiently in both the sf-9 (acnpv-permissive) and bmn (non-acnpv-permi ... | 1996 | 8709251 |
| codon usage in the prototype baculovirus--autographa californica nuclear polyhedrosis virus. | the overall codon usage profile of autographa californica nuclear polyhedrosis virus (acnpv) was analyzed, using uwgcg sequence analysis software package from the known protein coding gene sequences available in genbank release 72. the analysis revealed that although only 45% of the codon used by acnpv have g/c at wobble base position, 15 out of 20 acnpv codons over-utilized for their given amino acids has g/c at the wobble position indicating a possible selection of these codons. the difference ... | 1995 | 8714213 |
| assembly of virus-like particles in insect cells infected with a baculovirus containing a modified coat protein gene of potato leafroll luteovirus. | dna encoding the coat protein (p3) of a scottish isolate of potato leafroll virus (plrv) was inserted into the genome of autographa californica nucleopolyhedrovirus (acnpv) such that the coat protein was expressed either in an unmodified form or with the addition of the amino acid sequence mhhhhhhgddddkdamg at the n terminus (p3-6h). insect cells infected with these recombinant baculoviruses accumulated substantial amounts of p3 and p3-6h. p3 could not be recovered from cell extracts unless it w ... | 1996 | 8757974 |
| characterization of the lymantria dispar nucleopolyhedrovirus 25k fp gene. | the lymantria dispar nucleopolyhedrovirus (ldmnpv) gene encoding the 25k fp protein has been cloned and sequenced. the 25k fp gene codes for a 217 amino acid protein with a predicted molecular mass of 24870 da. expression of the 25k fp protein in a rabbit reticulocyte system generated a 27 kda protein, in close agreement with the molecular mass predicted from the nucleotide sequence. the gene is located between 40.3 and 40.8 map units on the viral genome. it is transcribed in a counterclockwise ... | 1996 | 8760443 |
| species-specific effects of the hcf-1 gene on baculovirus virulence. | the host cell-specific factor 1 gene (hcf-1) of the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) is required for the efficient replication and/or stability of reporter plasmids carrying an acmnpv-derived origin of dna replication in a cell-specific manner; hcf-1 is required for reporter plasmid replication or stability in tn-368 cells, a cell line derived from the cabbage looper trichoplusia ni, but not in iplb-sf-21 (sf-21) cells, a cell line derived from the fall army ... | 1996 | 8764020 |
| a common pathway for p10 and calyx proteins in progressive stages of polyhedron envelope assembly in acmnpv-infected spodoptera frugiperda larvae. | the assembly of the polyhedron envelope in baculovirus-infected cells has been the subject of several studies, yet it is still poorly understood. we have used immunogold-labelled antibodies to two baculovirus proteins, p10 and calyx (also referred to as polyhedron envelope protein or pep), to follow envelope assembly in acmnpv-infected tissues of spodoptera frugiperda larvae. we show that, in wild type virus, both proteins colocalize in fibrillar structures and associated electron-dense spacers ... | 1996 | 8774685 |
| green fluorescent protein as a tool for screening recombinant baculoviruses. | the gene encoding the green fluorescent protein (gfp) from the jellyfish aequorea victoria, ligated to the honeybee melittin signal peptide-encoding sequence, was inserted under transcriptional control of the polyhedrin promoter of the autographa californica nuclear polyhedrosis virus and expressed in the spodoptera frugiperda insect cell line sf9 during viral infection. the recombinant green fluorescent protein was identified by sds-page gel electrophoresis followed by coomassie blue staining o ... | 1996 | 8793839 |
| dynamic phosphorylation of autographa californica nuclear polyhedrosis virus pp31. | autographa californica nuclear polyhedrosis virus (acmnpv) pp31 is a nuclear phosphoprotein that accumulates in the virogenic stroma, which is the viral replication center in the infected-cell nucleus, binds to dna, and serves as a late expression factor. considering that reversible phosphorylation could influence its functional properties, we examined phosphorylation and dephosphorylation of pp31 in detail. our results showed that pp31 is posttranslationally phosphorylated by both cellular and ... | 1996 | 8794314 |
| initiation of baculovirus dna replication: early promoter regions can function as infection-dependent replicating sequences in a plasmid-based replication assay. | from the results of transient plasmid-based replication assays, it has been postulated that homologous regions (hrs) of autographa californica nuclear polyhedrosis virus (acmnpv) function as origins of viral dna replication. however, these assays vary in specificity according to the methodology used and may not be dependent solely on the presence of hr sequences. to determine the role that hrs and other sequences might play in the replication process, a series of plasmids containing specific del ... | 1996 | 8794340 |
| induction of apoptosis by baculovirus transactivator ie1. | apoptosis is induced upon infection of sf-21 cells by mutants of the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) lacking a functional p35 gene which encodes a stoichiometric inhibitor of members of the interleukin-1beta converting enzyme family of cysteine proteases (n.j. bump et al, science 269:1885-1888, 1995; r.j. clem, m. fechheimer, and l.k. miller, science 254:1388-1390, 1991). we found that transfection of sf-21 cells with the acmnpv ie-1 gene was sufficient to ... | 1996 | 8794358 |
| transcription, translation, and cellular localization of three autographa californica nuclear polyhedrosis virus structural proteins: odv-e18, odv-e35, and odv-ec27. | this paper identifies two structural proteins of the occluded derived viral envelope of autographa californica nuclear polyhedrosis virus (acmnpv): odv-e18 and odv-e35. in addition, we identify a protein, odv-ec27, that is incorporated into the capsid of occluded virus, which is not detected in budded virus. the genes for these proteins reside within the ie0 intron. the intron was sequenced, and five open reading frames (orf) were identified. orf 3 (genomic orf 143) codes for the odv envelope pr ... | 1996 | 8806491 |
| cfmnpv blocks acmnpv-induced apoptosis in a continuous midgut cell line. | morphological and molecular changes produced by autographa californica nuclear polyhedrosis virus (acmnpv) infection in a permissive cell line, iplb-sf-21ae (sf-21), of spodoptera frugiperda and a nonpermissive cell line, fpmi-cf-203 (cf-203), of choristoneura fumiferana are described. cf-203 cells inoculated with acmnpv showed a dna ladder and morphological changes such as plasma membrane granulation, blebbing, and nuclear fragmentation, which are characteristic of apoptosis. typical virus repl ... | 1996 | 8806500 |
| mapping functional domains in acmnpv pp31. | autographa californica nuclear polyhedrosis virus (acmnpv) replicates in the nucleus and produces a viral-modified form of the nuclear matrix called the virogenic stroma. the virogenic stroma is the site of viral dna packaging and nucleocapsid assembly and is thought to be the site of viral dna replication and rna transcription. acmnpv encodes a phosphoprotein, pp31, which localizes to the nucleus of uninfected insect cells and to the virogenic stroma of infected insect cells. pp31 has dna bindi ... | 1996 | 8806516 |
| identification and nucleotide sequence of an ecdysteroid udp-glucosyltransferase gene of spodoptera littoralis multicapsid nuclear polyhedrosis virus. | the spodoptera littoralis multicapsid nuclear polyhedrosis virus (slmnpv) is a member of the baculoviridae that shows a distant genetic relationship to the prototype autographa californica mnpv (acmnpv). using an acmnpv gene-specific probe, we identified and mapped an ecdysteroid udp-glucosyltransferase (egt) gene in the genome of slmnpv. sequence determination of a part from the hybridizing dna fragment revealed an open reading frame of 1548 nucleotides that exhibits 38% and 44% identity to the ... | 1995 | 8808334 |
| expression and analysis of a bacterial poly(hydroxyalkanoate) synthase in insect cells using a baculovirus system. | an improved method for expression of poly-beta-hydroxyalkanoate (pha) synthase from alcaligenes eutrophus has been developed using the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) in bti-tn-5b1-4 trichoplusia ni cells which results in high level production of active pha synthase. confirmation of expression of authentic pha synthase was obtained by western analysis which also revealed the presence of several apparent proteolytic cleavage products. n-terminal sequence dat ... | 1996 | 8812862 |
| establishment of a cell line derived from embryos of the potato tuber moth phthorimaea operculella (zeller). | a cell line from the main insect pest of potatoes in tropical and subtropical areas, phthorimaea operculella (zeller), was obtained from embryoculture. these cells were cultured in grace's modified medium. the cell line, designated ors-pop-93, had a heterogeneous population consisting of spherical and spindle cells with great capacity to adhere and a doubling time of 40 h. they were subcultured for more than 60 passages. their polypeptidic profile was different from profiles of other lepidoptera ... | 1995 | 8826086 |
| secretion of green fluorescent protein from recombinant baculovirus-infected insect cells. | trichoplusia ni (high five) and spodoptera frugiperda (sf21) cells were engineered for expression of epitope (flag)-tagged signal peptide-green fluorescent protein (gfp) fusions to examine the suitability of gfp as a secretory marker. the recombinant baculovirus-infected cells became fluorescent, and the high five cells but not sf21 cells secreted gfp in the culture medium as detected by the presence in the culture supernatant of a flag-immunoreactive 30-kda species and the characteristic 510-nm ... | 1996 | 8831686 |
| bifunctionality of the acmnpv homologous region sequence (hr1): enhancer and ori functions have different sequence requirements. | the autographa californica multinucleocapsid nuclear polyhedrosis virus (acmnpv) homologous region sequence hr1 is a putative origin of replication (ori) sequence and can also function as a transcriptional enhancer for delayed-early genes. we demonstrate that this 750-bp sequence, carrying five 28-bp core palin-dromes, enhances expression from the very late polyhedrin promoter up to 11-fold in a classical enhancer fashion in transient expression assays. enhancement is at the level of transcripti ... | 1996 | 8836032 |
| identification and characterization of an early gene in the lymantria dispar multinucleocapsid nuclear polyhedrosis virus. | the lymantria dispar multinucleocapsid nuclear polyhedrosis virus (ldmnpv) gene encoding g22 was cloned and sequenced. the g22 gene codes for a 191 amino acid protein with a predicted m(r) of 22,000. expression of g22 in a rabbit reticulocyte system generated a protein with an m(r) of 24,000, in close agreement with the molecular mass predicted from the nucleotide sequence. g22 is not significantly homologous to any known protein, nor is a g22 homologue present in the autographa californica mnpv ... | 1995 | 8847497 |
| temperature-sensitive mutations in the protein kinase-1 (pk-1) gene of the autographa californica nuclear polyhedrosis virus that block very late gene expression. | we have developed a genetic screen for temperature-sensitive mutations in the very late transcription apparatus of the autographa californica nuclear polyhedrosis virus. this method starts with the bacpaks virus, which has the escherichia coli lacz gene under the control of the very late polyhedrin promoter. the desired mutants are temperature-sensitive for beta-galactosidase production and can be complemented by wild-type virus, which lacks the lacz gene. two mutants created by nitrosoguanidine ... | 1996 | 8862393 |
| a baculovirus vector derived from immediately early gene promoter of autographa californica nuclear polyhedrosis virus. | a transfer vector was constructed in which the neomycin resistance (neo) gene was under the control of a copy of autographa californica nuclear polyhedrosis virus (acmnpv) ie1 gene promoter at the p10 locus. after cotransfection of spodoptera frugiperda (sf9) cells with the transfer vector and infectious acmnpv dna, the recombinant virus-containing neo gene was selected by serial passage of the mixed progenies from cotransfection. this was done at low moi in the presence of g418 in growth medium ... | 1996 | 8910650 |
| expression and loading of recombinant heavy and light chain homopolymers of rat liver ferritin. | the full-length genes for the heavy (h) and light (l) chains of ferritin isolated from a rat liver cdna library were amplified using polymerase chain reaction. each was inserted at the unique bglii site downstream of the p10 promoter of the baculovirus transfer vector pacuw21. the genes were transferred separately to infectious autographa californica nuclear polyhedrosis virus (acnpv) expression vectors after in vivo homologous recombination. ferritin homopolymers of either h or l chain were exp ... | 1996 | 8914851 |
| overexpression of actin in acmnpv-infected cells interferes with polyhedrin synthesis and polyhedra formation. | autographa californica m nuclear polyhedrosis virus (acmnpv) terminates host protein synthesis during the late stage of infection, at approximately 14 hr postinfection (hpi). if infection takes place in the presence of cytochalasin d (cd), however, host actin synthesis is transiently stimulated and continues to be synthesized until approximately 30 hpi, and the hyperexpression of polyhedrin is delayed from about 20 hpi until about 36 hpi (s. n. talhouk and l. e. volkman, virology 182, 626-634, 1 ... | 1996 | 8918923 |
| characterization of the ecdysteroid udp-glucosyltransferase gene from mamestra brassicae nucleopolyhedrovirus. | the ecdysteroid udp-glucosyltransferase (egt) gene of mamestra brassicae multinucleocapsid nucleopolyhedrovirus (mbmnpv) has been cloned and characterized. mbmnpv egt potentially encodes a protein of 528 amino acids. analysis of the substrate specificity of the mbmnpv egt protein showed that it mirrors that of autographa californica mnpv (acmnpv) egt. mbmnpv egt also appears to be secreted from infected cells. confirmation that the cloned gene encodes an active egt was obtained by transient expr ... | 1996 | 8922482 |
| gene transfer into hepatocytes and human liver tissue by baculovirus vectors. | gene therapy of liver diseases requires the development of efficient vectors for gene transfer in vivo. retroviral and adenoviral vectors have been shown to deliver genes efficiently into hepatocytes in vitro and in vivo. however, these vectors do not allow for exclusive infection of the liver which would be highly advantageous for in vivo gene therapy strategies. we have recently demonstrated that genetically modified baculoviruses (autographa californica nuclear polyhedrosis virus) efficiently ... | 1996 | 8930653 |
| the role of the acmnpv 25k gene, "fp25," in baculovirus polh and p10 expression. | a previous study showed that an autographa californica multicapsid nuclear polyhedrosis virus (acmnpv) 25k mutant produced less polyhedrin protein than wild-type (jarvis et al., j. virol. 66, 6903-6911, 1992). in this study, the role of the 25k gene product (acmnpv orf 61) in baculovirus gene expression was further investigated. five different viral 25k mutants expressed lower levels of polyhedrin protein and less cat activity under the control of the polh promoter compared to wild-type. polh rn ... | 1996 | 8941320 |
| effects of viral inhibitors of apoptosis in models of mammalian cell death. | apoptotic cell death is used as a defence against infection by viruses. to counter this protective mechanism, some viruses carry genes whose products can inhibit progression of the apoptotic process in the host cell. as it is clear that the core cell death mechanisms have been conserved through evolution, viral genes from various sources can be used to unravel these mechanisms in mammalian cells. we have produced transgenic mice that express the cowpox gene crma in their t cell compartment, and ... | 1996 | 8950470 |
| replication of simian virus 40 origin-containing dna during infection with a recombinant autographa californica multiple nuclear polyhedrosis virus expressing large t antigen. | autographica californica multiple nuclear polyhedrosis virus (acmnpv) has been shown to encode many of the enzymes involved in the replication of its own dna. although the acmnpv genome contains multiple sets of reiterated sequences that are thought to function as origins of dna replication, no initiator protein has yet been identified in the set of viral replication enzymes. in this study, the ability of a heterologous origin initiator system to promote dna replication in acmnpv-infected cells ... | 1997 | 8985377 |
| nucleotide sequence, insertional mutagenesis, and transcriptional mapping of a conserved region of the baculovirus autographa californica nuclear polyhedrosis virus (map unit 64.8-66.9). | the nucleotide sequence of a 2773-bp region of the autographa californica nuclear polyhedrosis virus hr3 variant (acmnpv) (map unit 64.4-68.2) has been determined. this region lies between the previously analyzed p6.9 and p80 genes. three open reading frames were contained within this region, potentially coding for proteins of 45, 40, and 12 kda. the 5' ends of transcripts capable of coding for the 45- and 12-kda proteins map to consensus baculovirus late transcription start sites (ataag), while ... | 1996 | 8989865 |
| role of early and late replication events in induction of apoptosis by baculoviruses. | autographa californica nuclear polyhedrosis virus (acmnpv) mutants that lack the apoptotic suppressor gene p35 cause apoptosis in spodoptera frugiperda sf21 cells. to identify a viral signal(s) that induces programmed cell death, we first defined the timing of apoptotic events during infection. activation of a p35-inhibitable caspase, intracellular fragmentation of host and acmnpv dna, and cell membrane blebbing coincided with the initiation of viral dna synthesis between 9 and 12 h after infect ... | 1997 | 8995679 |
| differential infectivity of two autographa californica nucleopolyhedrovirus mutants on three permissive cell lines is the result of lef-7 deletion. | we isolated two autographa californica nucleopolyhedrovirus (acmnpv) mutants that have infectivity similar to that of wild-type (wt) acmnpv in tn368 cells, but reduced budded virus and polyhedral inclusion body production in iplb-sf-21 and se1c cells. restriction endonuclease analysis and sequence analysis indicated that 3.2-kb (77.0-79.4 m.u.) and 4.4-kb (76.7-80.1 m.u.) regions, the location of four major open reading frames (orfs), pk2, orf-247, lef-7, and chitinase, were deleted in mutant t2 ... | 1997 | 9007061 |
| association of a baculovirus-encoded protein with the capsid basal region. | an open reading frame homologous to acmnpv orf9 (orf1629) was characterized in the orgyia pseudotsugata multinucleocapsid nuclear polyhedrosis virus (opmnpv). sequence analysis indicated that the opmnpv homolog (called orf2) encoded a protein predicted to contain 545 amino acids with a molecular weight of 61 kda. the first 80 amino acids did not have a counterpart in the acmnpv homolog. the remainder of the orf was poorly conserved with 29% amino acid identity overall with the acmnpv orf. howeve ... | 1997 | 9007067 |
| insecticidal activity of a recombinant baculovirus containing an antisense c-myc fragment. | attempts to develop baculovirus-based insecticides by insertion of genes encoding enzyme inhibitors, neuropeptides or toxins have met with some success. however, it is often difficult to ensure correct processing or secretion of the encoded peptides. here we tested a simpler strategy by insertion of an antisense fragment of a host gene to block translation of a protein essential for larval growth and development. we selected the c-myc gene for two main reasons: (i) its protein is known to be wel ... | 1997 | 9010314 |
| a new vector for efficient generation of p10-single-late-promoter recombinant baculoviruses. | a new baculovirus (bacten) was constructed in order to generate p10 recombinant expression vectors at high frequency. this virus is an acmnpv derivative, with the polyhedrin gene deleted and thus exhibiting p10 promoter as a single strong late promoter. the polyhedrin coding sequence was re-inserted subsequently under the control of the p10 promoter, in place of the p10 coding sequence. two flanking bsu36i restriction sites were inserted together with the polyhedrin coding region. bacten can, th ... | 1997 | 9015270 |
| transcriptional analysis and promoter activity of the spodoptera littoralis multicapsid nucleopolyhedrovirus ecdysteroid udp-glucosyltransferase gene. | the ecdysteroid udp-glucosyltransferase gene (egt) of spodoptera littoralis multicapsid nucleopolyhedrovirus (splimnpv) is a homologue of the autographa californica mnpv (acmnpv) egt gene, which has been found to block insect moulting. infection of larvae with an egt-deleted acmnpv resulted in enhanced mortality as compared to infection with the wild-type virus. consequently, deletion of an egt gene has been proposed as a tempting approach for enhancing the insecticidal properties of baculovirus ... | 1997 | 9018073 |
| autographa californica nuclear polyhedrosis virus p143 gene product is a dna-binding protein. | we have identified the protein product of the autographa californica nuclear polyhedrosis virus (acmnpv) p143 gene by constructing a recombinant baculovirus overexpressing the gene product p143. the overexpressed protein exhibited a relative mobility of approximately 140 kda and was stable for at least 12 hr after synthesis. immunoblotting using a monoclonal antibody developed against the overexpressed protein identified a similar polypeptide in acmnpv-infected cells which was detectable by 4 hr ... | 1997 | 9024814 |
| effects of diet-age and streptomycin on virulence of autographa californica m nucleopolyhedrovirus against the tobacco budworm. | addition of the antibiotic streptomycin to two artificial diets routinely used in bioassays of neonate lar vae of heliothis virescens (tobacco budworm) infected with autographa californica m nucleopolyhedrovirus (acmnpv) increased lethal times of the virus. after storage of diets for 3 weeks at 4 degrees c, lethal times of infected larvae were significantly slower compared to those for larvae bioassayed using diets stored for 2 weeks or less. the effect of diet-age on rate of mortality was not t ... | 1997 | 9028927 |
| expression of the mouse interleukin-2 receptor gamma chain in insect cells using a baculovirus expression vector--comparison with the human common gamma chain. | the gene encoding the gamma-chain of the mouse interleukin-2 receptor was expressed in lepidopteran insect cells using the baculovirus expression vector system. the corresponding gene was inserted under the polyhedrin promoter of the autographa californica nuclear polyhedrosis virus and expressed in the spodoptera frugiperda insect cell line sf9 during viral infection. the recombinant receptor protein was identified by immunoblotting in cell lysates prepared from insect cells infected with the p ... | 1997 | 9042425 |
| maturation of the dengue-2 virus ns1 protein in insect cells: effects of downstream ns2a sequences on baculovirus-expressed gene constructs. | a series of recombinant baculoviruses was constructed in order to study the influence of downstream ns2a sequences on the processing of the dengue virus ns1 glycoprotein in insect cells. ns1 alone was expressed at a high level in its native dimeric form and processed efficiently through the spodoptera frugiperda (sf) cell secretory pathway. recombinant ns1 was found associated with the plasma membrane and was also secreted into the extracellular medium. although both intra- and extracellular ns1 ... | 1995 | 9049346 |
| effect of methyltransferase inhibitors on the regulation of baculovirus protein synthesis. | in the presence of the methyltransferase inhibitor 3-deazaadenosine (3da-ado) the production of infectious autographa californica nuclear polyhedrosis virus (acmnpv) in tissue culture was only slightly affected, while the synthesis of very late proteins (polyhedrin and p10) was abolished. the synthesis of the influenza virus proteins ns1 and ha, expressed under the polyhedrin promoter, was also abolished by 3da-ado. furthermore, 3da-ado interfered with the shut-off of early and late acmnpv prote ... | 1995 | 9049353 |
| detection of antibodies against porcine parvovirus nonstructural protein ns1 may distinguish between vaccinated and infected pigs. | the humoral antibody response against the nonstructural protein ns1 and the structural protein vp2 of porcine parvovirus (ppv) was evaluated by immuno-peroxidase test (ipt) and enzyme linked immuno sorbent assay (elisa) using recombinant ppv antigens. the coding sequence for ns1 and vp2 was inserted into the baculovirus. autographa californica nuclear polyhedrosis virus (acnpv) genome resulting in two recombinant baculoviruses acnpv-ns1 and acnpv-vp2, respectively. sf9 cells (spodoptora frugidip ... | 1997 | 9050166 |
| enhanced bioactivity of recombinant baculoviruses expressing insect-specific spider toxins in lepidopteran crop pests | two genetically enhanced isolates of the autographa californica nuclear polyhedrosis virus (acmnpv) expressing insect-specific neurotoxin genes from the spiders diguetia canities and tegenaria agrestis were evaluated for their commercial potential. since prevention of feeding damage is of primary importance in assessing agronomic efficacy, a method for estimating the median time to cessation of feeding (ft50) was developed. neonate droplet feeding assays with preoccluded virus samples were condu ... | 1997 | 9056461 |
| polyomavirus major capsid protein vp1 is capable of packaging cellular dna when expressed in the baculovirus system. | using the p2bac dual multiple cloning site transfer vector, the polyomavirus major capsid protein gene vp1 was cloned for expression in the baculovirus-insect cell expression system. the 5-day-infected cellular lysate from this recombinant preparation was purified by cesium chloride density gradient centrifugation. capsid-like particles were observed in the resulting preparation. the purified particle preparation was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was s ... | 1997 | 9060642 |
| expression of rat steroid 5 alpha-reductase (isozyme-1) in spodoptera frugiperda, sf21, insect cells: expression of rat steroid 5 alpha-reductase. | the enzyme steroid 5 alpha-reductase (5 alpha r) catalyzes the reduction of testosterone (t) to 5 alpha-dihydrotestosterone (dht). in this study, the baculovirus expression system was used to overexpress rat 5 alpha r type i isozyme (r5 alpha r 1). the full length of r5 alpha r1 cdna was inserted into the autographa californica nuclear polyhedrosis virus (ac-mnpv) genome and expressed in spodoptera frugiperda, sf 21, insect cells. the expressed recombinant r5 alpha-r1 showed maximal enzymatic ac ... | 1997 | 9090798 |
| expression of foreign proteins on the surface of autographa californica nuclear polyhedrosis virus. | based on the method of direct cloning into the baculovirus genome by linearizing and re-ligation in presence of the target insert, we designed viral constructs that express foreign genes on the surface of baculovirus particles. we chose the glycosylated envelope protein gp41 of human immunodeficiency virus type 1 (hiv-1) as a model for displaying recombinant proteins on budded virus. the ectodomain of the envelope protein gp41 of hiv-1 was being fused to the entire baculovirus major coat protein ... | 1997 | 9105625 |
| n-terminal sequences from autographa californica nuclear polyhedrosis virus envelope proteins odv-e66 and odv-e25 are sufficient to direct reporter proteins to the nuclear envelope, intranuclear microvesicles and the envelope of occlusion derived virus. | baculovirus occlusion-derived virus (odv) derives its envelope from an intranuclear membrane source. n-terminal amino acid sequences of the autographa californica nuclear polyhedrosis virus (acmnpv) envelope proteins, odv-e66 and odv-e25 (23 and 24 amino acids, respectively) are highly hydrophobic. recombinant viruses that express the two n-terminal amino acid sequences fused to green fluorescent protein (23gfp or 24gfp) provided visual markers to follow protein transport and localization within ... | 1997 | 9108103 |
| organization and molecular characterization of genes in the polyhedrin region of the anagrapha falcifera multinucleocapsid npv. | a multinucleocapsid nuclear polyhedrosis virus (mnpv) isolated from the celery looper, anagrapha falcifera, has been proposed as a new virus based on differences in virulence and dna fragment profiles between this isolate and the autographa californica mnpv, the mnpv type species. in the present study, we examined the relatedness of the afmnpv and acmnpv genomes by (1) southern hybridization, (2) comparison of their genetic organization in the polyhedrin gene region (acmnpv ecori-i fragment), an ... | 1997 | 9125047 |
| baculovirus-mediated trans-epithelial transport of proteins in infected caterpillars. | baculovirus-mediated abnormal protein trafficking was studied in infected caterpillars by using heterologous proteins. the gene for human complement c1r was expressed in larvae of mamestra brassicae by a recombinant autographa californica nuclear polyhedrosis virus (acmnpv) vector. by following the time-course of recombinant c1r distribution among various tissues, cell types and cell organelles, we concluded that the dominant site of recombinant protein synthesis was the fat body, although some ... | 1997 | 9129671 |
| comprehensive physical map of the cydia pomonella granulovirus genome and sequence analysis of the granulin gene region. | a cloned strain of cydia pomonella granulovirus, cpgv-m1, was obtained using successive rounds of an in vivo limiting dilution method. a detailed physical map of the genome was constructed using 11 restriction enzymes. the region containing the granulin gene and an open reading frame immediately upstream of the granulin gene was sequenced. this region showed a high degree of homology to the equivalent region from cryptophlebia leucotreta granulovirus with 98% amino acid identity for the granulin ... | 1997 | 9129672 |
| baculovirus immediate-early promoter-mediated expression of the zeocin resistance gene for use as a dominant selectable marker in dipteran and lepidopteran insect cell lines. | the antibiotic zeocin, a derivative of phleomycin, was evaluated for use as a selection system in both dipteran and lepidopteran insect cell lines. growth of drosophila cell lines, kc1 and sl2, was inhibited at zeocin concentrations of 50 and 75 microg/ml, respectively, while the spodoptera cell line, sf9, was inhibited at a concentration of 250 microg/ml zeocin. the mammalian cytomegalovirus (cmv) and simian virus 40 (sv40) early promoters did not function in these insect cell lines. several ba ... | 1997 | 9133590 |
| late promoter selection in the baculovirus gp64 envelope fusion protein gene. | the upstream promoter region of the autographa californica multicapsid nuclear polyhedrosis virus (acmnpv) gp64 gene contains five copies of taag, the conserved sequence found at the transcriptional initiation sites of almost all baculovirus late genes. in acmnpv-infected sf9 cells, late transcription initiation is detected from only two upstream taag sites and not from three downstream taag sites. to examine several models for preferential taag site utilization, we constructed a series of recom ... | 1997 | 9168879 |
| loading of iron into recombinant rat liver ferritin heteropolymers by ceruloplasmin. | we have reported previously that the heavy chain of ferritin is required for iron incorporation by ceruloplasmin (j.-h. guo, m. abedi, and s. d. aust (1996) arch. biochem. biophys. 335(1)). the purpose of this study was to determine how many heavy chains were required for ceruloplasmin to interact with ferritin such that iron loading occurred. the cdna sequences encoding the heavy and light chains of rat liver ferritin were cloned into the baculovirus transfer vector pa-cuw51 under the control o ... | 1997 | 9169016 |
| genetically variable triplet repeats in a ring-finger orf of helicoverpa species baculoviruses. | nucleotide sequence analysis of the helicoverpa zea s-type nucleopolyhedrovirus (hzsnpv) genomic interval between the polh and iel genes has revealed an open reading frame (hoar orf) that contains a complex a 1-t rich triplet repeat region (rat-repeats). hoar orf is predicted to encode an acidic, arginine residue rich. 712 aa protein, with a c3hc4 (ring-finger) zinc binding motif. rat-repeats, distributed over 450 bp. consist of gat. aat, and gtt codons, correspond to asp, asn and val residues w ... | 1997 | 9178498 |
| expression of lacz reporter gene under the control of the polyhedrin promoter of spodoptera litura nuclear polyhedrosis virus. | promoter function of the putative polyhedrin-encoding gene (polh) of spodoptera litura nuclear polyhedrosis virus (s1mnpv) was determined by transferring it to the autographa californica nuclear polyhedrosis virus (acmnpv) through the acnpv polh based vector, pvl1393. three transfer vectors pcbt2, pcbt3 and pcbt4 were constructed by substituting the promoter and the neighbouring sequences of acnpv in pvl1393 by that of s1npv. the escherichia coli lacz gene was placed downstream from the s1npv po ... | 1997 | 9185860 |
| heterologous promoter recognition leading to high-level expression of cloned foreign genes in bombyx mori cell lines and larvae. | the baculovirus expression system using the autographa californica nuclear polyhedrosis virus (acnpv) has been extensively utilized for high-level expression of cloned foreign genes, driven by the strong viral promoters of polyhedrin (polh) and p10 encoding genes. a parallel system using bombyx mori nuclear polyhedrosis virus (bmnpv) is much less exploited because the choice and variety of bmnpv-based transfer vectors are limited. using a transient expression assay, we have demonstrated here tha ... | 1997 | 9185865 |
| identification and characterization of a baculovirus structural protein, vp1054, required for nucleocapsid formation. | the defect in a temperature-sensitive mutant of autographa californica nuclear polyhedrosis virus, tsn1054, was mapped and characterized. at the nonpermissive temperature of 33 degrees c, this mutant fails to form plaques upon infection of sf-21 cultured insect cells; infection is limited to a single cell, even though the infection proceeds through the very late phase. marker rescue mapping and dna sequencing identified the gene, orf 54, which was altered by a single nucleotide substitution in t ... | 1997 | 9188569 |
| a mechanism for negative gene regulation in autographa californica multinucleocapsid nuclear polyhedrosis virus. | the autographa californica multinucleocapsid nuclear polyhedrosis virus (acmnpv) ie-1 gene product (ie-1) is thought to play a central role in stimulating early viral transcription. ie-1 has been demonstrated to activate several early viral gene promoters and to negatively regulate the promoters of two other acmnpv regulatory genes, ie-0 and ie-2. our results indicate that ie-1 negatively regulates the expression of certain genes by binding directly, or as part of a complex, to promoter regions ... | 1997 | 9188574 |
| dna replication promotes high-frequency homologous recombination during autographa californica multiple nuclear polyhedrosis virus infection. | the relative ease with which foreign genes can be incorporated into the genome of the baculovirus autographa californica nuclear polyhedrosis virus (acmnpv) indicates that a highly efficient recombinational process exists within infected cells. however, it is unclear whether this is due to marker transfer mediated by host cell enzymes or recombination events promoted by acmnpv itself. to address the latter possibility, a pair of inverted repeat is50 elements derived from the bacterial transposon ... | 1997 | 9191843 |
| baculovirus transactivator ie1 is functional in mammalian cells. | ie1 of autographa californica multicapsid nuclear polyhedrosis virus acts as a transactivator of several viral promoters in insect cells. transient expression assays indicate that ie1 is involved in the activation of the early promoter he65 in both insect tn-368 and mammalian bhk-21 cell lines. ie1 activation of the he65 promoter was compared with ie1 activation of the early 39k promoter. in contrast to the he65 promoter, the 39k promoter was not inducible by ie1 in bhk-21 cells. | 1997 | 9191949 |
| differential requirements of two insect cell lines for growth in serum-free medium. | the development of a serum-free medium that supports the growth of cells from a spodoptera frugiperda and a lymantria dispar cell line is reported. a yeast hydrolysate provided the b-vitamin complex, and a combination of a meat hydrolysate and tryptose provided most of the free amino acids required for cell growth. supplemental cystine and methionine were required to achieve maximum cell growth. the serum or serum replacements used in earlier formulations were replaced with commercial lipid prep ... | 1997 | 9201517 |
| a role for baculovirus gp41 in budded virus production. | insect cells infected with tsb1074, a temperature-sensitive mutant of autographa californica nuclear polyhedrosis virus, exhibit a "single-cell-infection" phenotype whereby the infection progresses through the very late phase culminating in occlusion body formation, but neighboring cells do not become infected. marker rescue mapping and dna sequencing correlated a single nucleotide substitution within the baculovirus gp41 gene with the temperature-sensitive phenotype of tsb1074. the product of t ... | 1997 | 9217053 |
| complete sequence and transposon mutagenesis of the bamhi j fragment of cydia pomonella granulosis virus. | the bamhi j fragment of cydia pomonella granulosis virus was subcloned and subjected to transposon mutagenesis in escherichia coli using a tn3 derivative. after screening by restriction endonuclease digestion and polymerase chain reaction (pcr), 44 clones were selected representing insertions every 100 to 300 bp. the complete sequence was compiled and the transposon insertion sites mapped precisely by sequencing. analysis of the sequence revealed the presence of 7 potential open reading frames ( ... | 1997 | 9237352 |