Conjugation of DNA with protein using His-tag chemistry and its application to the aptamer-based detection system.

We propose a novel method to prepare a DNA-protein conjugate using histidine-tag (His-tag) chemistry. Oligo-DNA was modified with nitrilotriacetate (NTA), which has high affinity to a His-tag on recombinant protein via the complexation of Ni(2+). Investigations using a microplate which displayed a complementary DNA-strand revealed that a NTA-modified DNA-protein conjugate was formed and immobilized in the presence of Ni(2+) on the microplate. We then adopted alkaline phosphatase (AP) as a model protein, and application of the DNA-AP conjugate was demonstrated in a thrombin aptamer-based detection system with a detection limit of approximately 10 nM.