Baculovirus expression system

A method for producing a recombinant baculovirus of which the genome comprises:

• one or more transgene(s) each encoding a protein maturation enzyme, and n transgenes each encoding a polypeptide of interest to be expressed at a different locus, said method comprising the steps of:
• a) preparing, in an insect cell, a recombinant baculovirus genome capable of replicating which comprises said one or more transgene(s), each encoding a protein maturation enzyme, and said n transgenes each encoding a polypeptide of interest, by homologous recombination between:
• a1) a replication deficient baculovirus genome in which n genes essential for viral replication located in different regions of the viral genome are non-functional and which comprises said one or more transgene(s), each encoding a protein maturation enzyme, and
• a2) n transfer vectors each comprising:
• i) a nucleotide sequence enabling to restore the function of one of the n non-functional genes essential for viral replication located in different regions of the viral genome,
• ii) one of said n transgenes encoding a polypeptide of interest, each polypeptide of interest to be expressed at a different locus,
• the set of nucleotide sequences i) of the n transfer vectors being capable of restoring the replication of the replication deficient baculovirus genome,
• n being an integer at least equal to 2;
• and
• b) generating a recombinant baculovirus in an insect cell which comprises the recombinant baculovirus genome obtained at step a),
• wherein said homologous recombination between the replication deficient baculovirus genome and the n transfer vectors takes place in a single transfection step in the insect cell.