The invention relates to recombinant novirhabdoviruses having at least one sequence encoding a polypeptide of interest added to their genome. Said recombinant novirhabdoviruses are useful as gene vectors, for producing recombinant proteins, or for producing vaccines for fish or for higher vertebrates.
1. A recombinant DNA construct consisting of:
a) a transcriptional termination/polyadenylation sequence of novirhabdovirus M gene as set forth in the nucleotide sequence of SEQ ID NO: 4;
b) a transcription initiation sequence of novirhabdovirus G gene as set forth in the nucleotide sequence of SEQ ID NO: 5;
c) a non-transcribed intergenic dinucleotide of a novirhabdovirus preceded by said transcriptional termination/polyadenylation sequence, or followed by said transcription initiation sequence; and
d) an open reading frame encoding a protein of interest.
2. An antigenomic cDNA of the genome of a novirhabdovirus, comprising one or more recombinant DNA constructs claim 1 , inserted in said cDNA between a stop codon of a first endogenous ORF and a start codon of a second endogenous ORF of the host novirhabdovirus.
3. A recombinant novirhabdovirus containing a genomic RNA complementary to the antigenomic cDNA of claim 2 .
4. An antigenomic cDNA of the genome of a novirhabdovirus, comprising three recombinant DNA constructs of claim 1 , inserted in said cDNA between a stop codon of a first endogenous ORF and a start codon of a second endogenous ORF of the host novirhabdovirus.
5. A recombinant novirhabdovirus containing a genomic RNA complementary to the antigenomic cDNA of claim 4 .
6. The recombinant novirhabdovirus of claim 5 , wherein the recombinant novirhabdovirus is a recombinant Infectious Hematopoietic Necrosis Virus (IHNV).
7. A method of producing a protein of interest comprising expressing the recombinant novirhabdovirus of claim 3 in vertebrate cells in culture.
8. A live attenuated vaccine comprising the recombinant novirahbdovirus of claim 3 .
9. An antigen delivery system for vaccination of a bird or mammal comprising the recombinant novirhabdovirus of claim 3 .
10. A method for expressing an antigenic protein of interest in vitro in a low temperature expression system, comprising expressing a recombinant of IHNV or Viral Harmorrhagic Septicemia Virus (VHSV) comprising at least one recombinant DNA construct of claim 1 , wherein the open reading frame encoding a protein of interest encodes said antigenic protein.
11. A low temperature in vitro expression system, wherein said expression system comprises a recombinant IHNV or VHSV, comprising at least one recombinant DNA construct of claim 1 , wherein the open reading frame encoding a protein of interest encodes an antigenic protein of interest, and a vertebrate cell susceptible of infection by said recombinant virus and grows at low temperature.
12. A method for expressing in vitro an antigenic protein of interest, comprising:
a) infecting a vertebrate cell susceptible of infection by IHNV or VHSV and which grows at low temperature with a recombinant IHNV or VHSV comprising at least one recombinant DNA construct of claim 1 , wherein the open reading frame encoding a protein of interest encodes an antigenic protein of interest;
b) culturing said cell at a temperature of about 14° C. to about 20° C.; and
c) recovering the antigenic protein of interest produced by said cell.
13. An antigenomic cDNA of the genome of a novirhabdovirus, comprising two recombinant DNA constructs of claim 1 , inserted in said cDNA between a stop codon of a first endogenous ORF and a start codon of a second endogenous ORF of the host novirhabdovirus.