Keywords

Phage Display System, Zearalenone, Mimotope, Enterokinase

Abstract

Abstract It is planned to use p¢ø phage display system to express Zearalenone (ZEN) mimotope peptide and verify its reactogenicity. The expression vector pC89-CZEN was constructed by ligating the oligonucleotide containing the ZEN mimotope sequence and the enterokinase cleavage site with the pC89S4 phagemid that had been double digested by EcoR I and BamH I. pC89-CZEN was transformed into competent XL1-Blue to obtain the engineered strain pC89-ek-zen, which was then superinfected with KM13 and induced by IPTG for expression. After purification, phage particles displaying zearalenone mimotopes were obtained. The bacterial concentration OD600 during KM13 superinfection, the bacterial concentration OD600 during IPTG induction, the amount of IPTG added, and the induction expression temperature and time were optimized to explore the best expression conditions; the reactogenicity was determined by ELISA and compared before and after enterokinase digestion. Binding effect of mimotope peptides and ZEN antibodies. The results showed that the expression vector pC89-CZEN was successfully constructed. The optimal expression conditions were that the bacterial concentration OD600 was 0.4 during KM13 superinfection, the bacterial concentration OD600 during IPTG induction was 0.6, and the added amount of IPTG was the final concentration of 1.0 mmol/L. The expression temperature was 24¡ãC and the time was 8 hours. The binding effect after enzyme digestion was significantly higher than before enzyme digestion.

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Original research done by Liu Renrong, Xu Fuyong, Xu Ling, Zhu Lixin, Qiu Xuemei

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