The Preparation and Application of NHXFS1 Polyclonal Antibody

Abstract: NHXFS1 was a novel powerful vacuolar Na+/H+ antiporter gene which was obtained from Arabidopsis thaliana antiporter gene AtNHX1， Oryza sativa antiporter gene OsNHX1 and Dendranthema morifolium antiporter gene DmNHX1 by DNA family shuffling technology. In this study， we prepared and identified the polyclonal antibody against the C-terminal fragment of NHXFS1. For expression and purification of NHXFS1-C terminal protein， the expression vector pET32a-NHXFS1 antigen containing 6×His tag was constructed and transferred into the E.coli strain BL21 (DE3). After IPTG induced， the fusion protein was purified by Ni-NTA superflow cartridges and was immunized with rabbits for polyclonal antibody preparation. The ELISA results showed that the potency of polyclonal antibody reached 1:128 000. To identify the antibody specificity， the vacuolar was extracted from the yeast stain expressing NHXFS1 and OsNHX1 protein and analyzed by Western blot. The result indicated that we have obtained high specificity polyclonal antibody that could be used for further study of Na+/H+ antiporter and the molecular mechanism of plant salinity.

CSTR: 32200.14.cjcb.2012.05.0009