Abstract：
:An inducible in vitro cell culture system was developed to assay HCV replication by direct biochemical means. A transcription plasmid containing a T7 promoter at the 5' end, full-length cDNA of the HCV genome, a ribozyme sequence from the antigenomic strand of hepatitis delta virus and a T7 terminator was prepared. To facilitate high-level transcription of HCV RNA, HepG2 cells were infected with replication deficient adenovirus containing the T7 RNA polymerase gene and later transfected with the transcription plasmid containing the full-length HCV genome. This transfection-based cell culture system expressed high levels of HCV structural (core, El and E2) and non-structural proteins (NS3 and NS5B) detectable by Western blot and immunofluorescence assays. Production of HCV RNA transcripts and presence of replicative negative strand of HCV was confirmed by ribonuclease protection assay indicating replication of HCV in the transfected HepG2 cell. The transfected HepG2 cells assembled 50-60 nm virus-like particles, which could be aggregated by anti-E2 antibodies. This model can be utilized for studying mechanisms of HCV replication, assembly of HCV particles and to test potential anti-HCV compounds.
journal_name
J Virol Methodsjournal_title
Journal of virological methodsauthors
Myung J,Khalap N,Kalkeri G,Garry R,Dash Sdoi
10.1016/s0166-0934(01)00278-6subject
Has Abstractpub_date
2001-05-01 00:00:00pages
55-67issue
1-2eissn
0166-0934issn
1879-0984pii
S0166093401002786journal_volume
94pub_type
杂志文章abstract：:The appearance of cytomegalovirus (CMV) proteins in infected fibroblasts was determined by flow cytometry. The sequential production of immediate early (IE), early (E), and late (L) proteins reacting with respective monoclonal antibodies (mAbs) E13, 58/5, and 24/4 was determined in fibroblasts infected with the AD-169...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(93)90105-z
更新日期：1993-12-15 00:00:00
abstract：:Varicella zoster virus (VZV) PCR is highly sensitive compared to traditional detection methods like culture and direct fluorescent antibody testing (DFA); however, the high cost of commercial assays prohibits their use in many clinical laboratories. Major contributors to cost are the nucleic acid extraction and the PC...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.02.009
更新日期：2014-06-01 00:00:00
abstract：:With the approval of the first HIV-1 integrase inhibitor raltegravir and a second one in phase III clinical development (elvitegravir), genotypic and phenotypic resistance assays are required to guide antiretroviral therapy and to investigate treatment failure. In this study, a genotypic and phenotypic recombinant vir...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2009.06.015
更新日期：2009-11-01 00:00:00
abstract：:The disruption of the viral coat of human immunodeficiency virus by Triton X-100, a nonionic detergent, is a time-dependent process which requires incubation times of 30 min or longer. Conditions for the production of a noninfectious sample from a viral pellet that can be used to measure reverse transcriptase activity...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(97)00169-9
更新日期：1998-01-01 00:00:00
abstract：:Sensitive, precise and rapid detection tests are needed in the quality control of rabies vaccine for rabies virus nucleoprotein. Previous studies for quantitation of rabies virus nucleoprotein focused on enzyme-linked immunosorbent assay (ELISA). A novel immunoassay for rapid determination of rabies virus nucleoprotei...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.06.002
更新日期：2014-09-01 00:00:00
abstract：:TaqMan Mismatch Amplification Mutation Assay (TaqMAMA) is a highly sensitive allelic discrimination method. The mismatch amplification mutation assay (MAMA) is based on preferential amplification of mutant allele by the 'MAMA' primer, which is designed to have two mismatches with the wild-type allele and only one mism...
journal_title：Journal of virological methods
pub_type： 杂志文章,随机对照试验
doi：10.1016/j.jviromet.2008.07.020
更新日期：2008-11-01 00:00:00
abstract：:HIV-1 integrase (HIV-1 IN), a well-validated antiviral drug target, catalyzes multistep reactions to incorporate viral DNA into the genome of the host cell; these include both a 3'-processing (3'P) reaction and a strand transfer reaction. These enzymatic activities can be measured in vitro with short DNA oligonucleoti...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2012.05.003
更新日期：2012-09-01 00:00:00
abstract：:The World Health Organization recommends screening donor blood for HIV in centralized laboratories. This recommendation contributes to quality, but presents specimen transport challenges for resource-limited settings which may be relieved by using dried blood spots (DBS). In sub-Saharan Africa, most countries screen d...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2011.07.001
更新日期：2012-01-01 00:00:00
abstract：:A method is described for detecting enteroviruses in both freshwater and marine sediments. Viruses were recovered from sediments by elution into 6% beef extract at pH 9.0 and concentration with polyethylene glycol 6000. The recovery efficiency ranged from 6 to 55% for marine sediments and 16 to 77% for freshwater sedi...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(85)90101-6
更新日期：1985-02-01 00:00:00
abstract：:A long RT-PCR (LRP) protocol was developed recently for robust amplification of a near full-length HCV genomic sequence from clinical samples, followed by efficient cloning [Fan, X., Xu, Y., Di Biceglie, A.M., 2006. Efficient amplification and cloning of near full-length hepatitis C virus genome from clinical samples....
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2006.12.002
更新日期：2007-05-01 00:00:00
abstract：:Combining primers created from the sequence information of two baculo-like viruses of penaeid shrimp, Baculovirus penaei (BP) and Monodon baculovirus (MBV), produced a 750 bp band on a 0.8% agarose gel using White Spot Syndrome Virus (WSSV), from Penaeus monodon, as the DNA template. The PCR fragment was ligated to a ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(96)02128-3
更新日期：1997-01-01 00:00:00
abstract：:Different viruses belonging to the genus Vesivirus infect a broad range of animals, and cause gastroenteritis, vesicular skin lesions, hemorrhagic disease, respiratory diseases and other conditions. A recent report on Vesivirus viremia, as detected by PCR, in samples from patients with hepatitis of unknown etiology in...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2008.12.008
更新日期：2009-04-01 00:00:00
abstract：:Sequence analysis of duck plague virus (DPV) revealed that there was a 528bp (B fragment) deletion within the UL2 gene of DPV attenuated vaccine strain in comparison with field virulent strains. The finding of gene deletion provides a potential differentiation test between DPV virulent strain and attenuated strain bas...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2017.08.021
更新日期：2017-11-01 00:00:00
abstract：:Double-stranded RNA (dsRNA) is in many cases the only available template for molecular and diagnostic studies of RNA viruses. A novel mycovirus with a five dsRNAs segmented-genome served as a model system for the amplification and cloning of dsRNA segments using several PCR-based methods. Sequences obtained by the cla...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2010.08.013
更新日期：2010-11-01 00:00:00
abstract：:Grapevine shoot cultures infected by Grapevine vitivirus A (GVA) were grown on Quorin-Lepoivre basic medium and submitted to in vitro chemotherapy and thermotherapy sanitation techniques. Ribavirin (Rb) at 20gml(-1), dihydroxypropyladenine (DHPA) at 60gml(-1) and their combination (RbDH) were added to the proliferatin...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2007.06.008
更新日期：2007-12-01 00:00:00
abstract：:Cell-mediated cytotoxicity against cytomegalovirus (CMV)-infected fibroblasts (FS-4 cells) was investigated by a non-radioactive assay, and by DNA fragmentation ELISA and LDH release assay and the assays were compared to the standard chromium release assay. Fragmentation of DNA and LDH activity were detected in the su...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(95)01954-5
更新日期：1996-01-01 00:00:00
abstract：:The role of environmental reservoirs in avian influenza virus (AIV) transmission has been investigated during AIV-associated outbreaks. To date, no method has been defined for detection of AIV from mud samples. A procedure using elution and polyethylene glycol (PEG) concentration steps was designed to detect AIV by RT...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2011.10.013
更新日期：2012-01-01 00:00:00
abstract：:Accurate detection and quantitation of viruses can be beneficial to plant-virus interaction studies. In this study, three SYBR green real-time RT-PCR assays were developed to quantitate grapevine leafroll-associated virus 3 (GLRaV-3) in infected vines. Three genomic regions (ORF1a, coat protein and 3'UTR) were targete...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.09.022
更新日期：2014-12-15 00:00:00
abstract：:Prunus necrotic ringspot ilarvirus (PNRSV) exists as a number of biologically distinct variants which differ in host specificity, serology, and pathology. Previous nucleotide sequence alignment and phylogenetic analysis of cloned reverse transcription-polymerase chain reaction (RT-PCR) products of several biologically...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(99)00051-8
更新日期：1999-07-01 00:00:00
abstract：:Receptor-mediated endocytosis was employed for a highly efficient transport of oligodeoxynucleotides into hepatoma cell line PLC/PRF/5. The oligodeoxynucleotides were bound to the asialofetuin-poly-L-lysine conjugate and this complex was internalized by the cells via asialoglycoprotein receptor, an endocytic receptor ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(93)90181-p
更新日期：1993-04-01 00:00:00
abstract：:Detection of classical swine fever virus (CSFV) and its discrimination from other pestiviruses can be achieved by virus isolation (VI) in cell cultures, antigen detection, or molecular analysis. To simplify the latter, a 5'-nuclease assay (TaqMan) was developed for the rapid and specific detection of CSFV with the min...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(97)00208-5
更新日期：1998-06-01 00:00:00
abstract：:A fluorescent DNA probe (DV2.P1) specific to the conserved distal 3'-noncoding region (nucleotides 10653-10678) of dengue 2 virus and a pair of flanking primers (DV2.L1 and DV2.U2) were designed to formulate a dengue 2-specific fluorogenic polymerase chain reaction (PCR). In addition, DV2.L1 was also used as a reverse...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(99)00166-4
更新日期：2000-04-01 00:00:00
abstract：:A simple and fast sap-direct RT-PCR (reverse transcription-polymerase chain reaction) for the rapid detection of 3 viroids of the genus Coleviroid is presented. The templates for cDNA synthesis were obtained directly from the sap of coleus using a pipettor, a common tool in molecular biology laboratories, and 3 coleus...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2011.03.018
更新日期：2011-06-01 00:00:00
abstract：:Nucleic-acid amplification technology (NAT) assays have been implemented for HCV and HIV-1 in the United States, and many parts of Europe, Australia and Asia. Nucleic acid detection assays utilize many different technologies, and the WHO International Standards for nucleic acid tests are widely used to compare them. C...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2004.01.021
更新日期：2004-06-01 00:00:00
abstract：:Bovine herpesvirus 4 (BoHV-4) is a gamma herpesvirus with no clear disease association. Previous studies have demonstrated that macrophages can harbour persistent BoHV-4. Since mesenchymal stem cells in bone marrow regulate the differentiation and proliferation of adjacent haematopoietic precursors, such as macrophage...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2005.02.019
更新日期：2005-08-01 00:00:00
abstract：:The nucleocapsid (N) protein of a coronavirus plays a crucial role in virus assembly and in its RNA transcription. It is important to characterize a virus at the nucleotide level to discover the virus's genomic sequence variations and similarities relative to other viruses that could have an impact on the functions of...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2019.113806
更新日期：2020-03-01 00:00:00
abstract：:Identification of African swine fever virus (ASFV) proteins recognised by cytotoxic T lymphocytes (CTL) from swine surviving ASFV/NH/P68 infection was assessed using expression vectors based on the Pseudomonas aeruginosa outer membrane lipoprotein I gene (oprI). Viral antigens expressed as fusion lipoproteins were sho...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/s0166-0934(98)00105-0
更新日期：1998-11-01 00:00:00
abstract：:A general strategy for the construction of an internal standard for the polymerase chain reaction (PCR) is described together with its application in the evaluation of clinical samples. This internal standard is a plasmid containing a modified target sequence that is co-amplified with the native target using the same ...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(92)90099-y
更新日期：1992-09-01 00:00:00
abstract：:Concurrent infection of pigs with two or more pathogens is common in pigs under intensive rearing conditions. Porcine circovirus type 2 (PCV2), porcine parvovirus (PPV), porcine reproductive and respiratory syndrome virus (PRRSV), classical swine fever virus (CSFV), Japanese encephalitis virus (JEV) and pseudorabies v...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/j.jviromet.2014.06.027
更新日期：2014-11-01 00:00:00
abstract：:A logistic ('growth curve') model is formulated and applied to the relationship between numbers of infections (local lesions) produced by a virus on inoculated plants and the concentration of the virus in the inoculum. This model has the advantages of being simple, data-based and therefore not founded on limiting post...
journal_title：Journal of virological methods
pub_type： 杂志文章
doi：10.1016/0166-0934(87)90084-x
更新日期：1987-12-01 00:00:00
