Deoxyribonucleotide (DNA) was successfully extracted in a few hours by reverse micelles, which were formed and functionalized by a cationic quaternary ammonium surfactant in isooctane. Several parameters which affect the DNA transfer from an aqueous phase to a reverse micellar phase, such as the natures of a surfactant and a solvent, salt concentration in a feed solution, and hydrophobicity of micelles, were investigated. The use of a cationic surfactant was found to yield a high transfer rate of DNA into reverse micelles. We have attributed the DNA transfer to the electrostatic interaction between the cationic surfactant and the negatively charged DNA. We found that the higher the hydrophobicity of a surfactant used to form micelles, the better the DNA transfer efficiency in the forward extraction. Preferential extraction of DNA was achieved by using twin long-alkyl chains of cationic surfactants. However, pH in the aqueous feed solution did not affect the extraction performance of reverse micelles. On the other hand, alcohol, as a co-solvent plays an important role in the back extraction of DNAs. Complete back-extraction of DNAs was achieved without any conformational change by adding an appropriate alcohol to the reverse micellar phase.