Helicase-dependent amplification (HDA) is based on natural DNA replication mechanisms. Initially, the coordinated action of helicases unwinds and separates the template DNA duplex. The primer can hybridize with the free single-stranded templates, and the subsequent extension by a DNA polymerase will result in DNA amplification (Figure 6(A) ). The original reaction reported in the literature is performed at 37 °C for the entire process, and more than a million-fold amplification of DNA fragments can be achieved from nanogram quantities of genomic DNA [79] . Unlike the PCR, HDA uses helicases instead of heat, thus eliminating the need for any denaturation steps. Nevertheless, two additional accessory proteins are required in this approach: MutL to stimulate helicase unwinding activity and a single-strand binding (SSB) protein to prevent premature re-association of the separated ssDNA. A thermostable helicase may be also advantageous for HDA. Recently, a new helicase was developed from Thermoanaerobacter tengcongensis, which can be operated at temperatures from 45 °C to 65 °C [80] , so HDA reactions are now generally performed at the higher temperature of 65 °C. The use of thermostable helicase led researchers to abandon both the MutL and SSB proteins, while simultaneously improving the DNA yield of the reaction [81] . This simple thermal management option makes HDA very attractive for the development of simple portable DNA diagnostic devices and point-of-care testing.