Prokaryotic expression, purification of Tldi1 protein and preparation of its polyclonal antibody in Salmonella Typhimurium.

This study aimed to construct a prokaryotic expression vector for the T6SS immunity protein Tldi1 of Salmonella Typhimurium and to prepare its specific polyclonal antibody, thereby facilitating further functional research on Tldi1.The dominant antigenic sequence of Tldi1 was identified through bioinformatics analysis. The target gene was amplified from the genomic DNA of Salmonella Typhimurium SL1344. The recombinant vector pET-32a-tldi1 was then constructed and transformed into Escherichia coli. The recombinant Tldi1 protein was expressed following IPTG induction and purified by nickel affinity chromatography. KM mice were immunized with the purified protein to generate polyclonal antibodies, and the antibody titer and specificity were then determined. The recombinant expression vector was successfully constructed, and the purified Tldi1 protein achieved a purity of over 90%. The prepared polyclonal antibody demonstrated a high titer of 1:256,000and strong specificity. This study successfully obtained high-activity Tldi1 protein and its specific antibody, which provide essential experimental materials for analyzing the T6SS-mediated interbacterial competition mechanism and for establishing a rapid immunological detection method for Salmonella.