Chemistry: molecular biology and microbiology – Virus or bacteriophage – except for viral vector or...
Patent
1990-11-28
1994-05-10
Schwartz, Richard A.
Chemistry: molecular biology and microbiology
Virus or bacteriophage, except for viral vector or...
4353201, 4352402, 536 2372, 536 242, 535 32, C12N 701, C12N 1539, C12N 1586, C12N 510
Patent
active
053106717
DESCRIPTION:
BRIEF SUMMARY
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention is in the field of recombinant DNA technology and relates to fowlpox virus as a vector for foreign DNA.
2. Description of the Prior Art
Several viruses with DNA genomes have been used to carry and express genes from other viruses or other species. Viruses used in such a way are known as `vectors` and genes, other than their own, expressed in such a way are referred to as `foreign genes`. One of the primary requirements for a virus to be used as a vector in this manner is a suitable site for insertion of the foreign gene. If insertion of a gene into a site in the virus causes disruption of some function essential for growth, then such a site could not be used. Suitable sites are those at which an insertion does not disrupt any functions or those whose functions are not essential for viral growth and therefore can be disrupted with impunity. Such sites are known as `non-essential regions`. The phrase `non-essential` in this context means non-essential for growth under at least some conditions in which the virus can be grown in vitro and under at least some conditions in which it survives in vivo.
Examples of viruses which have been used as vectors by virtue of the fact that they contain non-essential regions are orthopoxviruses, adenoviruses and herpesviruses, although the regions used may be different in each case. Vaccinia virus (VV), which has been used as a vector, is an orthopoxvirus, a member of the pox virus family. Fowlpoxvirus (FPV), the subject of this patent application, is also a pox virus, but is a member of a different genus, the avipoxviruses. VV can be grown in tissue culture, and foreign genes can be inserted into the viral genome during this process. Several regions of VV have been found to be non-essential in vitro in more than one tissue-culture system. These include: large regions towards the left hand end, B. Moss et al., J. Virology 40, 387-395, (1981) who describe a mutant VV having a deletion 6.4 megaDaltons from the left-hand end; D. Panicali et al., J. Virology 37 1000-1010, 1981) who describe a mutant VV having a deletion starting 6.85 megaDaltons from the left-hand end; the thymidine kinase (TK) gene, D. Panicali and E. Paoletti, Proc. Natl. Acad. Sci. USA 79, 4927-4931 (1982); M. Mackett et al., J. Gen. Virol. 45, 683-701, (1982); and the vaccinia growth factor (VGF) gene, R. M. L. Buller it al., J. Virology 62, 866-974 (1988). Sites such as these might also be non-essential for growth in vivo. However in the case of both the TK and VGF gene, it has been found that although growth in tissue culture is unaffected or only slightly affected by insertion into the gene, growth and virulence in vivo are markedly affected, R. M. L. Buller el al., Nature 317, 813-815 (1985) and loc. cit. (1988), showing that these genes are not completely non-essential for growth of the virus in vivo. This in vivo attenuation may however be useful if it reduces unwanted pathogenic effects of the virus and accordingly growth in vivo with accompanying attenuation is a valid growth condition for the purpose of this invention.
Some sites are essential in some tissue culture systems and non-essential in others. For example there is a gene in VV which is essential for replication in human cells but which is non-essential in chicken embryo fibroblast cells, S. Gillard et al., Proc. Natl. Acad. Sci. USA 83, 5573-5577 (1986). Another gene in the related orthopoxvirus cowpox virus is essential for growth in chinese hamster ovary cells but not for growth in chicken embryo fibroblast cells, D. Spehner et al., J. Virology 62, 1297-1304 (1988). These examples show that differences in the tissue culture systems can affect which regions are non-essential.
The VV genome is far from being completely mapped and relatively little has been published about the FPV genome. It is known that FPV has a TK gene which has about 60% homology with the VV TK gene at the amino acid level , D. B. Boyle et al., Virology 156, 355-365 (1987). Like the VV TK gene, it ser
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Binns Matthew M.
Boursnell Michael E. G.
Campbell Joan I. A.
Tomley Fiona M.
British Technology Group Limited
Mosher May E.
Schwartz Richard A.
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