摘要：Salmonella enterica serovar typhimurium is one of the important worldwide health
issues. The hilA gene encodes a transcriptional activator that regulates expression of
the majority of the genes responsible for the Salmonella invasive phenotype. In the
present study, PCR was performed using our designed primers for amplification of
upstream and downstream regions of Salmonella typhimurium hilA gene. Each DNA
fragment was T/A-cloned into pGEM-T easy vector and then sub-cloned into pET32
expression vector together with Kanamycin resistance gene. The recombinant plasmid
was transformed into bacteria (Salmonella typhimurium) using electroporation. The
hilA-Knockout mutant was characterized to evaluate the predicted role of the hilA gene
in virulence. Quantitative RT-PCR was carried out to study the impact of hilA knock
out on the expression of downstream genes including invF and invA. We confirmed
the successful preparation of hilA gene construct (pET32-up-kan-down) followed by
the efficient electroporation of the construct to bacterial cells. The homologous
recombination resulted in the hilA confirmed by PCR. We demonstrated that hilA
knockout leads to attenuation of invA and invF genes expression. The hilA knockout
strain may be useful for the development of efficient vaccines against the Salmonella
typhimurium.
