Selection of recombinant vaccinia viruses on the basis of plaque formation
We developed a procedure for isolation of recombinant vaccinia viruses (re-VV) based solely on plaque formation, without a requirement for specific cell lines, selective medium or special staining. The system consists of two components (i) a mutant non-plaque-forming VV and (ii) a plasmid vector that, through homologous recombination, can simultaneously introduce a foreign gene and repair the mutation in the VV genome. The mutant VV contains a deletion of the vp37 gene, encoding a 37-kDa protein component of the viral outer envelope that is required for efficient viral spread on cell monolayers. The plasmid vector contains a functional vp37, a strong synthetic VV early/late promoter, unique restriction sites for gene insertion, and flanking segments of VV DNA for homologous recombination. Following infection and transfection of cells with the mutant VV and plasmid vector, respectively, re-VV are identified and isolated by their ability to form plaques. To evaluate the system, a re-VV that expresses the gene encoding influenza virus hemagglutinin (HA) was isolated simply by picking visible plaques. © 1995.
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Elsevier
1995
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Online Access: | http://hdl.handle.net/20.500.12792/3907 http://hdl.handle.net/10261/294482 |
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dig-inia-es-10261-2944822023-02-20T10:39:09Z Selection of recombinant vaccinia viruses on the basis of plaque formation Blasco Lozano, Rafael Moss, B. We developed a procedure for isolation of recombinant vaccinia viruses (re-VV) based solely on plaque formation, without a requirement for specific cell lines, selective medium or special staining. The system consists of two components (i) a mutant non-plaque-forming VV and (ii) a plasmid vector that, through homologous recombination, can simultaneously introduce a foreign gene and repair the mutation in the VV genome. The mutant VV contains a deletion of the vp37 gene, encoding a 37-kDa protein component of the viral outer envelope that is required for efficient viral spread on cell monolayers. The plasmid vector contains a functional vp37, a strong synthetic VV early/late promoter, unique restriction sites for gene insertion, and flanking segments of VV DNA for homologous recombination. Following infection and transfection of cells with the mutant VV and plasmid vector, respectively, re-VV are identified and isolated by their ability to form plaques. To evaluate the system, a re-VV that expresses the gene encoding influenza virus hemagglutinin (HA) was isolated simply by picking visible plaques. © 1995. 2023-02-20T10:39:09Z 2023-02-20T10:39:09Z 1995 journal article Gene 158(2): 157-162 (1995) 0378-1119 http://hdl.handle.net/20.500.12792/3907 http://hdl.handle.net/10261/294482 10.1016/0378-1119(95)00149-Z en none Elsevier |
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We developed a procedure for isolation of recombinant vaccinia viruses (re-VV) based solely on plaque formation, without a requirement for specific cell lines, selective medium or special staining. The system consists of two components (i) a mutant non-plaque-forming VV and (ii) a plasmid vector that, through homologous recombination, can simultaneously introduce a foreign gene and repair the mutation in the VV genome. The mutant VV contains a deletion of the vp37 gene, encoding a 37-kDa protein component of the viral outer envelope that is required for efficient viral spread on cell monolayers. The plasmid vector contains a functional vp37, a strong synthetic VV early/late promoter, unique restriction sites for gene insertion, and flanking segments of VV DNA for homologous recombination. Following infection and transfection of cells with the mutant VV and plasmid vector, respectively, re-VV are identified and isolated by their ability to form plaques. To evaluate the system, a re-VV that expresses the gene encoding influenza virus hemagglutinin (HA) was isolated simply by picking visible plaques. © 1995. |
format |
journal article |
author |
Blasco Lozano, Rafael Moss, B. |
spellingShingle |
Blasco Lozano, Rafael Moss, B. Selection of recombinant vaccinia viruses on the basis of plaque formation |
author_facet |
Blasco Lozano, Rafael Moss, B. |
author_sort |
Blasco Lozano, Rafael |
title |
Selection of recombinant vaccinia viruses on the basis of plaque formation |
title_short |
Selection of recombinant vaccinia viruses on the basis of plaque formation |
title_full |
Selection of recombinant vaccinia viruses on the basis of plaque formation |
title_fullStr |
Selection of recombinant vaccinia viruses on the basis of plaque formation |
title_full_unstemmed |
Selection of recombinant vaccinia viruses on the basis of plaque formation |
title_sort |
selection of recombinant vaccinia viruses on the basis of plaque formation |
publisher |
Elsevier |
publishDate |
1995 |
url |
http://hdl.handle.net/20.500.12792/3907 http://hdl.handle.net/10261/294482 |
work_keys_str_mv |
AT blascolozanorafael selectionofrecombinantvacciniavirusesonthebasisofplaqueformation AT mossb selectionofrecombinantvacciniavirusesonthebasisofplaqueformation |
_version_ |
1767603620478451712 |