Recombinant modified vaccinia ankara (MVA) vaccinia virus containing restructured insertion sites
AssigneeThe United States of America, as represented by the Secretary, Department of Health and Human Services
InventorsBernard Moss, Linda S. Wyatt, Patricia L. Earl
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Abstract
The present invention relates to recombinant modified vaccinia Ankara (MVA) virus containing restructured sites useful for the integration of heterologous nucleic acid sequences into an intergenic region (IGR) of the virus genome, where the IGR is located between two adjacent, essential open reading frames (ORFs) of the vaccinia virus genome, wherein the adjacent essential ORFs are non-adjacent in a parental MVA virus used to construct the recombinant MVA virus, and to related nucleic acid constructs useful for inserting heterologous DNA into the genome of a vaccinia virus, and further to the use of the disclosed viruses as a medicine or vaccine.
Claims20
A recombinant modified vaccinia Ankara (MVA) virus comprising a heterologous nucleic acid sequence between two adjacent essential open reading frames (ORFs) of the MVA virus genome, wherein the MVA virus lacks non-essential ORFs that are present between the corresponding two essential ORFs in the parental MVA virus.
A method for producing a stable, recombinant modified vaccinia Ankara (MVA) virus, the method comprising:
- (a) transfecting a cell with a nucleic acid construct comprising;
- (i) a first nucleic acid sequence comprising an at least 100 contiguous polynucleotide region that is at least 75% identical to an at least 100 contiguous polynucleotide region from a first essential MVA virus ORF;
- and
- (ii) a second nucleic acid sequence comprising an at least 100 contiguous polynucleotide region that is at least 75% identical to an at least 100 contiguous polynucleotide region from a second essential MVA virus ORF;
- wherein the first and second essential MVA virus ORFs are separated by at least one non-essential ORF in the MVA virus genome, and wherein the first and second nucleic acid sequences are adjacent to each other in the isolated nucleic acid construct;
- (b) infecting the transfected cell with a MVA virus;
- (c) culturing the infected cell under conditions suitable to allow homologous recombination between the nucleic acid construct and the MVA virus genome;
- and
- (d) isolating the recombinant MVA virus.
Dependencies are inferred from the claim text.