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Stojanovic and coworkers created DNA nanomachines called “DNA spiders” on various path patterns constructed on DNA origami (Figure 1.11b) [87]. The DNA spiders consisted of three DNA strand legs and one capture DNA strand (capture leg). The three legs of the DNA spiders each contained a DNA enzyme (DNAzyme leg) that could hydrolyze RNA. Single‐stranded DNAs containing a cleavable RNA site were arranged on DNA origami as a track for the DNA spider to walk. The DNA spider was immobilized at a specific position on the DNA origami using a trapping DNA strand, followed by dissociation and initiation of walking on the track. The DNA spider bound to the ssDNAs on DNA origami, cleaved the RNA site in the ssDNAs using its DNAzyme function, and walked autonomously. The DNA spider moved forward along the predetermined track and finally stopped at a specific point on the DNA origami where ssDNAs did not have an RNA site for cleavage. All of these processes, including initiation, walking, and stopping, were controlled in a programmed manner. In addition, by measuring the position of the DNA spider on DNA origami using super‐resolution microscopy, it was found that the spider moved at a speed of 3 nm/min.

Figure 1.11 Assembly line with a DNA walker capturing gold particles and DNA spider molecule walking in a track on a DNA origami. (a) The DNA walker binds to the DNA strand on the DNA origami with three legs, and gold particles (AuNPs) are collected with three hands.

Source: Gu et al. [86]/with permission of Springer Nature.

The DNA walker stops at three places on DNA origami and receives AuNPs (C1–C3) to be transferred by rotating PX‐JX₂ DNA devices. Multiple operation on DNA origami and corresponding AFM image. (b) The DNA spider binds onto the DNA origami using three legs hybridized to ssDNAs (cleavage site contains RNA) in the track. DNAzyme for cleavage of RNA site in the ssDNA is introduced to three legs.

Source: Lund et al. [87]/with permission of Springer Nature.

DNA strands in the track before and after cutting (gray and light gray circles) and stopping DNA strands (right side circles). The path for walking with instruction (start, follow, turn, and stop) can be programmed on the DNA origami. AFM image of DNA spider molecule walking on the DNA origami track. Start (top), walking (middle), and stop (bottom). (c) A DNA motor system created on the DNA origami. A motor‐track (gray ssDNAs) was constructed on the DNA origami and the movement of the DNA motor (black ssDNA) was examined.

Source: Wickham et al. [88]/with permission of Springer Nature.

Stepwise movement of a DNA motor observed in real time by high‐speed AFM.