关键词: 单增李斯特菌, prsA1, 生物信息学, 原核表达, 多克隆抗体

Abstract: Listeria monocytogenes is a common food-borne pathogen that causes listeriosis and poses a tremendous threat to food safety. prsA1 is conservative and specific. Signal peptides, transmembrane regions and spatial antigen epitopes of PrsA1 were predicted in this study by SignalP 4.1 Server program, TMHMM Server V.2.0 program and SEPPA 2.0 program, respectively. The results showed that the N terminus of PrsA1 contained a signal peptide and a transmembrane region, and that the protein had good epitopes. Therefore, it could be used as a detection target. On this basis, non-transmembrane region sequence of prsA1, Δ84prsA1, was acquired through PCR. The recombinant plasmid pET30a-Δ84prsA1 was constructed and transferred into Escherichia coli to induce expression of Δ28PrsA1. Δ28PrsA1 was purified by Ni-IDA-Sefinose Column. The titer of polyclonal antibody prepared by purified Δ28PrsA1 was determined by indirect ELISA, and the titer was as high as 1∶128 000. Western blotting analysis showed that the polyclonal antibody could recognize PrsA1 protein extracted from Listeria monocytogenes. In this study, bioinformatics methods was used to screen the detection target and analyze the epitopes, and polyclonal antibody was prepared successfully, which provided a practical basis for the screening of detection target and immunological detection of Listeria monocytogenes.

Key words: Listeria monocytogenes, prsA1, bioinformatics, prokaryotic expression, polyclonal antibody