A functionality of on/off switching to regulate conductivity should be among the prerequisites for the utilization of DNA duplex as a conducting material. We have reported that variation of temperatures can regulate the triplex formation and dissociation, thereby regulating charge transport in DNA. Based on this result, the on/off switching of conductivity at dual triplex forming sites in DNA was attempted, using two types of oligonucleotide (ODNs) with different strand lengths. Variation of temperatures produced DNA duplexes without any triplex site and with single and double triplex sites, by which the intramolecular charge transport in DNA duplex could be regulated at the respective triplex-forming sites. This may promote the utility of DNA duplex as a potential constituent of electronic nanodevices.