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?:abstract
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Objective: Prokaryotic expression of affinity peptides to spike protein of transmissible gastroenteritis virus (TGEV) and analysis of viral affinity activity were conducted to provide basis for establishment of TGEV diagnostic methods Method: In this study, a tandem gene containing the TGEV spike protein affinity peptide was synthesized After sub-cloning, the gene was inserted into prokaryotic expression vectors of pET-32a and pGEX-6p-1 to obtain recombinant plasmids pET-32a-SQHT and pGEX-6p-SQHT The recombinant plasmids were identified by single enzyme digestion of BamH I, double enzyme digestion of BamH I/Xho I and sequencing Then, the recombinant plasmids were transformed into E coli Rosetta (DE3) and induced by IPTG to obtain the expression of recombinant protein Finally, the biological activity of expression products was measured Result: A gene about 150 bp of TGEV spike protein affinity peptide was obtained by sub-cloning Two recombinant plasmids, pET-32a-SQHT and pGEX-6p-SQHT, were constructed and expressed by induction successfully The molecular weights of the two recombinant proteins were 25 and 31 ku, respectively Western blot analysis showed that the two recombinant proteins had good affinity with TGEV virions Dot-ELISA analysis showed that the minimum binding titer of the two recombinant proteins binding to the TGEV virions was TCID sub 50 /sub 5x10 sup 2 /sup mL sup -1 /sup Specificity experiments showed that the recombinant proteins TRX-SQHT and GST SQHT did not bind to PEDV or PoRV
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