ArticleDNA and cell biology2009
Generation of recombinant vaccinia viruses via green fluorescent protein selection.
Article in DNA and cell biology, 2009. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 10 papers.
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Who cites it
10 citing papers in PubMed, 13 citations in OpenAlex.
- The Multiplicity of Infection of Recombinant Vaccinia Virus Expressing the T7 RNA Polymerase Determines the Rescue Efficiency of Vesicular Stomatitis Virus.Frontiers in microbiology · 2022Article
- Cytotoxic effect of dual fluorescent-labeled oncolytic herpes simplex virus type 1 on mouse tumorigenic cell lines.Research in pharmaceutical sciences · 2019Article
- Genomic Characterization ofViruses · 2019Article
- Construction of Various γ34.5 Deleted Fluorescent-Expressing Oncolytic herpes Simplex type 1 (oHSV) for Generation and Isolation of HSV-Based VectorsIranian biomedical journal · 2017Article
- Investigation of IRES Insertion into the Genome of Recombinant MVA as a Translation Enhancer in the Context of Transcript Decapping.PloS one · 2015Article
- Vaccinia reporter viruses for quantifying viral function at all stages of gene expression.Journal of visualized experiments : JoVE · 2014Article
- Vaccinia virus induces rapid necrosis in keratinocytes by a STAT3-dependent mechanism.PloS one · 2014Article
- Generation of recombinant Orf virus using an enhanced green fluorescent protein reporter gene as a selectable marker.BMC veterinary research · 2011Article
- Construction and characterisation of a recombinant fowlpox virus that expresses the human papilloma virus L1 protein.Journal of translational medicine · 2011Article
- Development of Vaccinia reporter viruses for rapid, high content analysis of viral function at all stages of gene expression.Antiviral research · 2011Article
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6 authors at 2 institutions in 1 country.
Funding
Abstract
We developed a rapid method to generate recombinant vaccinia viruses (rVVs) based upon a bicistronic cassette encoding the gene for green fluorescent protein (GFP) and a foreign gene of interest separated by an internal ribosome entry site (IRES). As proof-of-concept, we inserted a mutant env gene of human immunodeficiency virus (HIV) into the cassette, which was cloned into the vaccinia virus (VV) insertion vector pSC59 under the control of the early-late VV synthetic promoter and flanked by disrupted tk gene sequences. To generate rVVs, 293T cells were inoculated with wild-type (wt) VV, followed by transfection of the modified pSC59 vector containing the bicistronic cassette, which allows expression of GFP and the protein of interest. Next, GFP-positive cells were isolated by flow cytometry or by picking under a fluorescent microscope. Thymidine kinase-deficient (Tk(-)) 143B cells were then exposed to lysates of GFP-positive 293T cells and cultured in the presence of bromodeoxyuridine. This selection allows only Tk(-) rVV to remain viable. We demonstrated the success of this GFP selection strategy by expressing high levels of mutant HIV Env. Our approach shortens the time needed to generate rVVs and represents a practical approach to generate recombinant proteins.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.