Compositions and methods for recombinant parvovirus production
AssigneeAavnergene Inc.
InventorsQizhao Wang, Daozhan Yu, Yu Liu, Cuiping Chen, Rongze Yang
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Abstract
Expression constructs and methods for recombinant parvovirus production are disclosed. In some embodiments, the expression construct encodes (1) adenovirus E4 and E2a proteins, (2) parvovirus proteins necessary for the production of the recombinant parvovirus, and (3) a recombinant parvovirus genome, thus allowing production of the recombinant parvovirus by transfecting a host cell with a single expression construct.
Claims20
A single polynucleotide expression construct for production of a recombinant parvovirus, the expression construct comprising:
- (1) a modified adenovirus E4 coding region that encodes E4 open reading frame 6/7;
- (2) a modified adenovirus E2a coding region that encodes an E2a protein;
- (3) an adenovirus VA RNA coding region that encodes adenovirus VAI RNA and VAII RNA;
- (4) a parvovirus protein coding region that encodes parvovirus proteins necessary for the production of the recombinant parvovirus;
- (5) a recombinant parvovirus sequence comprising (i) an expression cassette comprising a nucleotide sequence encoding a transgene and a regulatory element operably linked to the nucleotide sequence, and (ii) at least one ITR at one end of the expression cassette;
- and
- (6) one or more regulatory elements that allow expression of the E4 open reading frame 6/7, the E2a protein, the adenovirus VAI RNA and VAII RNA, and the parvovirus proteins necessary for the production of the recombinant parvovirus in a host cell,
- wherein the single polynucleotide expression construct has a size of 7-25 kb.
A single polynucleotide expression construct for production of recombinant parvovirus, the expression construct comprising:
- (a) a modified adenovirus E4 coding region encoding adenovirus E4orf6/7, wherein the modified adenovirus E4 coding region comprises (i) partial or complete deletion of E4orf6/7 intron 1, or (ii) a partial or complete deletion of E4orf6/7 intron 2, or both (i) and (ii);
- (b) a modified adenovirus E2a coding region encoding an adenovirus E2a protein, wherein the modified adenovirus E2a coding region comprises at least one deletion in early-E2a intron 1, or later-E2a intron 1, or E2a intron 2;
- (c) a sequence encoding one or more adenovirus VA RNAs;
- and
- (d) one or more regulatory elements operably linked to (a), (b) and (c),
- wherein the single polynucleotide expression construct has a size of 2-11 kb.
Dependencies are inferred from the claim text.