Detection and comparison of protein-DNA interactions using DNA-BIND plate and horseradish peroxidase-based colorimetric assay |
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| Authors: | Liu Bin Qin Yu Wang Jinzhong Wang Ying |
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| Institution: | aTEDA School of Biological Sciences and Biotechnology, Nankai University, TEDA, Tianjin 300457, People’s Republic of China;bCollege of Basic Medical Science, Tianjin Medical University, Tianjin 300070, People’s Republic of China;cKey Laboratory of Molecular Microbiology and Technology, Ministry of Education, TEDA, Tianjin 300457, People’s Republic of China;dTianjin Key Laboratory of Microbial Functional Genomics, TEDA, Tianjin 300457, People’s Republic of China |
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| Abstract: | We describe a procedure for detection and comparison of protein–DNA interactions using DNA–BIND plate and horseradish peroxidase (HRP)-based colorimetric assay. Amino-modified oligonucleotide was covalently immobilized on the surface of DNA–BIND plate. After the complementary oligonucleotide was annealed, the plate was incubated with protein to allow sequence-specific DNA binding. Primary antibody and HRP-labeled secondary antibody were then employed, and colorimetric assay was conducted before the absorbance was read. This is a sensitive, specific, and high-throughput method that has been applied not only in the detection of protein–DNA interaction but also in the quantitative comparison of DNA-binding capabilities among wild-type and mutant proteins. |
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