[Prokaryotic expression and antibody preparation of human GALNT3-sol protein]

In order to detect the expression of GALNT3 in various tumor tissues, the prokaryotic expression vector of human GALNT3-sol (a truncation of GALNT3 being deleted of the hydrophobic trans-membrane domain) was constructed, and then the recombinant GALNT3-sol protein was expressed and purified from E.coli, followed by the preparation of polyclonal antibody against GALNT3-sol and characterization of its properties.

The human cDNA of GALNT3-sol (1 755 bp)was amplified from MKN45 cell line and cloned into expression vector pET5b/GALNT3-sol, then transformed into E.coli BL21(DE3), in which the GALNT3-sol protein was induced by IPTG and then purified by Electrophoresis.Mice were immunized with the purified protein and the anti-serum was collected at different time intervals.Properties of the anti-serum were further detected by ELISA and Western blot.

The prokaryotic expression vector of pET15b/GALNT3-sol was constructed successfully.Human GALNT3-sol protein was expressed in E.coli after IPTG induction.The titer of the obtained anti-serum reached 1:25 600, and its specificity was proved by Western blot.

Human GALNT3-sol protein can be successfully expressed in E.coli, and the specific anti-human GALNT3-sol antibody can be obtained by immunization of mice, which makes it possible to further investigate the role of GALNT3 in the progression of various tumors.