Seleção, caracterização e clonagem dos genes fljB e groEL agonistas dos receptores de reconhecimento de padrão do sistema imune das aves

Abstract

The recombinant production of innate immune system pattern recognition receptor agonists has provided a new tool for the production of immunostimulants for mammals. The molecular pattern associated to the pathogen (PAMP), flageline, coded by the fljB gene of the S. typhimurium and the molecular pattern associated to the damage (DAMP), HSP60, coded by the groEL gene of the S. typhimuriumand S. enteritidis, are recognized by pattern recognition receptors (PRRs) of bird innate immune system. In the present study, we performed the cloning of genetic fragments of genes fljB from the S. typhimurium and groEL from the S. typhimurium and S. enteritidis inserted in expression vector pET100/D-TOPO and transformed in E. coli TOP10 cells. The clones were evaluated by colony PCR, plasmidial DNA PCR and genome sequencing in order to confirm the presence of these genes. In the colony PCR, we identified the presence of the genes groEL (S. enteritidis), groEL (S. enteritidis) and fljB (S. typhimurium) in 80%, 60% and 80% of the transformed colonies, respectively. The cloning system adopted allowed the production of clones identical to the HSP60 genetic fragments and flaneline of the Salmonella strains, allowing the posterior use of these clones in gene expression trials, with the future potential of being used as non-specific immunostimulant for birds.