The study of DNA-binding proteins is crucial in understanding gene regulatory networks. We developed a new method for the enrichment of DNA-binding proteins based on the variability of DNA-protein complexes' solubility in different ionic strength solutions. 0.14M sodium chloride was determined as the most efficient extraction concentration to precipitate DNA-binding proteins. SDS-PAGE analysis revealed that some high-abundance proteins were removed effectively and at the same time DNA-binding proteins were isolated in this simple process. Twenty kinds of proteins were identified in the acquired sample by 1-D gel-LC-MS/MS. Furthermore, computerized analysis of MS data showed that quite a number of unmatched peptides have the classic structure of leucine zipper or zinc finger, which were symbolic elements of transcription factors. These results suggested that this new method can acquire DNA-binding proteins effectively and allow improvement in the isolation of high-quality DNA-binding proteins.

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Superoxide dismutase (SOD) activity is an important measure of plant stress tolerance used in cultivar improvement. At present, we are unaware of any widely available immunological reagents for the detection of SOD in Oryza sativa (common Asian rice) or other plants. In this study, we used insilico B-cell epitope prediction tools to generate peptides which were immunized into rabbits to yield polyclonal antibodies against Cu/Zn SODs. Immunoblotting demonstrated that the antibody specifically recognized both native and denatured Cu/Zn SODs in rice. In addition, this antibody can confirm the expression tendency of endogenous OsCu/Zn SODs under heat stress by immunoblotting, and has a positive reaction in tomato leaf extracts, as well as human Hela cells. Chloroplast content of Cu/Zn SODs in rice can be identified by ELISA indirect competition method using this antibody. These results suggest that this Cu/Zn SOD rabbit polyclonal antibody may be a useful tool for elucidating the biological functions of Cu/Zn SODs in plants.