EXPRESSION OF GST-SNAIL FUSION PROTEIN IN PROKARYOTIC CELLS AND PREPARATION OF POLYCLONAL ANTIBODY AGAINST SNAIL / 解剖学报

ABSTRACT

Objective The aim of this study was to prepare anti-snail polyclonal antibody and make it widely useful in snail detection. Methods The DNA fragment encoding human full length 264 amino acid of snail was obtained by PCR from cDNA library of human umbilical vein epithelial cells and was cloned into pGEX-4T-1 plasmid expressing glutathione S-transferase(GST).The GST-snail fusion protein was expressed by E.coli BL21 after IPTG induction and purified from total proteins of BL21 transformed by the recombinant plasmid pGEX-4T-1/snail.The New Zealand rabbit was immunized with the purified fusion protein to prepare polyclonal antiserum.The antiserum was identified by western blotting and immunofluorescent staining. Results The prokaryotic expression plasmid pGEX-4T-1/ snail was successfully constructed,and the fusion protein GST-snail was expressed efficiently.The polyclonal antibody raised in the rabbit could react specifically with snail in human cells.Conclusion The snail antiserum was of good purity with high titer and specificity which could satisfy the requirement for studying immuno-analysis on snail protein.