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Titolo:
Novel bovine lentiviral vectors based on Jembrana disease virus
Autore:
Metharom, P; Takyar, S; Xia, HH; Ellem, KAO; Macmillan, J; Shepherd, RW; Wilcox, GE; Wei, MQ;
Indirizzi:
Royal Childrens Hosp, SASVRC, Gene Therapy Unit, Brisbane, Qld 4029, Australia Royal Childrens Hosp Brisbane Qld Australia 4029 ane, Qld 4029, Australia Royal Childrens Hosp, Queensland Clin Genet Serv, Brisbane, Qld 4029, Australia Royal Childrens Hosp Brisbane Qld Australia 4029 ane, Qld 4029, Australia Royal Childrens Hosp, Queensland Inst Med Res, Brisbane, Qld 4029, Australia Royal Childrens Hosp Brisbane Qld Australia 4029 ane, Qld 4029, Australia Univ Queensland, Royal Childrens Hosp, Dept Paediat & Child Hlth, Brisbane, Qld, Australia Univ Queensland Brisbane Qld Australia ld Hlth, Brisbane, Qld, Australia Murdoch Univ, Div Vet & Biomed Studies, Perth, WA, Australia Murdoch UnivPerth WA Australia t & Biomed Studies, Perth, WA, Australia
Titolo Testata:
JOURNAL OF GENE MEDICINE
fascicolo: 3, volume: 2, anno: 2000,
pagine: 176 - 185
SICI:
1099-498X(200005/06)2:3<176:NBLVBO>2.0.ZU;2-E
Fonte:
ISI
Lingua:
ENG
Soggetto:
HUMAN-IMMUNODEFICIENCY-VIRUS; SUSTAINED GENE-EXPRESSION; NONDIVIDING HUMAN-CELLS; RETROVIRAL VECTORS; EFFICIENT TRANSDUCTION; SEQUENCE-ANALYSIS; SKIN FIBROBLASTS; BONE-MARROW; HIV; LONG;
Keywords:
Jembrana disease virus; bovine lentiviral vector; gene transfer; gene expression;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
56
Recensione:
Indirizzi per estratti:
Indirizzo: Wei, MQ Royal Childrens Hosp, SASVRC, Gene Therapy Unit, Brisbane, Qld 4029, Australia Royal Childrens Hosp Brisbane Qld Australia 4029 4029, Australia
Citazione:
P. Metharom et al., "Novel bovine lentiviral vectors based on Jembrana disease virus", J GENE MED, 2(3), 2000, pp. 176-185

Abstract

Background Safety is a concern that must be addressed prior to any clinical use of human immunodeficiency virus (HIV)-based lentiviral vectors in human patients. Unfortunately, efforts to examine the biosafety of the vectorsin preclinical animal models are hampered due to the lack of animal modelsfor HIV infection. We have developed new lentiviral vectors based on the recently characterised Jembrana Disease Virus (JDV), which infects a specific species of cattle naturally in Ball, Indonesia. Methods Sequences from the JDV genome were amplified by splicing overlap extension polymerase chain reaction (PCR) for the construction of transfer vectors as well as a packaging construct. Co-transfection of these two plasmids into 293T cells with a third encoding a G glycoprotein of vesicular stomatitis virus produced pseudotyped, disabled, replication defective JDV vector particles. Viral titre was obtained by transducing the cells with the supernatant harvested from transfectants and determining the number of cellsexpressing the transgene. PCR and Southern blotting were used to detect the presence of potential replication-competent viruses as well as transgene integration. Results Bicistronic JDV vectors encoding the green fluorescent protein (GFP) and the neomycin phosphotransferase were harvested with a titre range of0.4-1.2 x 10(6) colony forming units/ml from vector-producing cells and were further concentrated by ultracentrifugation to the high titre of approximately 10(7) CFU/ml. Vectors encoding GFP were shown to transduce and integrate efficiently into the chromosomes of a range of primary and transformedcells of different origins in different differentiation status, including growth-arrested cells, with an efficiency of 25-75%. Exhaustive testing with a marker gene transfer assay in combination with a reverse transcriptase assay and PCR amplification of samples of serially passaged, transduced cells showed that no detectable amount of replication competent lentivirus (RCL) was produced. Conclusions We showed the feasibility of the development of gene transfer vectors based on a non-primate bovine lentivirus, which will provide the opportunity for examination of the efficacy and biosafety of lentiviral vector-mediated gene transfer in vivo in animal models. JDV-based vectors may beapplicable and more readily acceptable than those from HIV for human gene therapy. Copyright (C) 2000 John Wiley & Sons, Ltd.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 09/07/20 alle ore 14:03:10