Portal de Periódicos da CAPES

Artigo Acesso aberto Produção Nacional Revisado por pares

BIBTEX RIS

Optimization of canine interleukin-12 production using a baculovirus insect cell expression system

2016; BioMed Central; Volume: 9; Issue: 1 Linguagem: Inglês

10.1186/s13104-016-1843-7

1756-0500

Cristiane Garboggini Melo de Pinheiro, Mayara de Oliveira Pedrosa, Naiara Carvalho Teixeira, Ana Paula Dinis Ano Bom, Monique M. van Oers, Geraldo Gileno de Sá Oliveira,

Transgenic Plants and Applications

Interleukin-12 is an important cytokine in mediating cellular immune responses.Recombinant single-chain canine IL-12 was produced in a baculovirus-insect cell system with the aim of conducting further studies on modulation of immune responses in dogs. To optimize the production of recombinant canine IL-12, a classical baculovirus and a modified vector (chitinase A and v-cathepsin knockout) were used containing a native or an optimized insert of canine IL-12. The optimized IL-12 construct contained the GP64 signal peptide and was synthesized with optimized codons for expression in Trichoplusia ni cells. Dot-blot and Western blot analysis showed the highest production levels of recombinant IL-12 protein by the use of the modified baculovirus vector containing the optimized insert, at a multiplicity of infection of five and at 48 h after infection. The recombinant cytokine was successfully purified and showed a good degree of purity, integrity, folding, and yield, with very little endotoxin contamination. Recombinant canine IL-12 induced IFN-γ in canine lymphocytes, indicating that it was biologically active.Therefore, this study describes an efficient method to produce adequate amounts of biologically active canine IL-12, useful for immunomodulation studies in dogs.