In Vitro Selection of a DNA Ligand that Selectively Binds to the Anticancer Agent Methotrexate

Abstract

Abstract A DNA ligand that selectively binds to the anticancer agent methotrexate was identified using the in vitro selection method. A pool of single-stranded DNAs consisting of random sequences was incubated with a methotrexate-immobilized agarose gel; bound DNAs were then collected and amplified using the polymerase chain reaction (PCR). This selection process was repeated six times. The selected DNAs were cloned and were found to contain two common sequences. One of these selected DNA clones selectively bound to the methotrexate-immobilized gel and showed a high level of discrimination against the gel immobilized with folic acid, which has a similar chemical structure to methotrexate. In the selection process, a buffer solution was used for DNA binding to the methotrexate-immobilized gel and water was used for elution from the gel. Ultraviolet absorption measurement showed that the conformation of the DNA ligand in the binding buffer was different from that in elution water.