Experimental Validation of Signal Dependent Operation in Whiplash PCR Ken Komiya 1 , Masayuki Yamamura 1 , and John A. Rose 2⋆ 1 Department of Computational Intelligence and Systems Science, Interdisciplinary Graduate School of Science and Engineering, Tokyo Institute of Technology, (komiya,my)@dis.titech.ac.jp 2 Institute of Information Communication Technology, Ritsumeikan Asia Pacific University, jarose@apu.ac.jp Abstract. Whiplash PCR (WPCR), which implements self-directed operation, programmed within a single DNA molecule, is a potential candidate for both mathematical and biological applications. However, WPCR-based methods are known to suffer from a serious efficiency problem called back-hybridization (BH). Previously, we proposed and partially validated a new rule-protect operation to abolish BH. In this work, we experimentally demonstrate the ability of rule- protect to drive multi-step WPCR. Successful implementation of isothermal op- eration at physiological temperatures is an essential benchmark for biological ap- plications. We also propose the use of rule-protect for external signalling to con- trol computational operation. Consequently, signal-dependent self-directed oper- ation, which is conceptually new to DNA computing, is achieved. The present architecture, provided with sensing ability, allows a composite system design layering computational reactions, and would be suitable for functioning as the central processing unit of this system. 1 Introduction In early studies in DNA-based computing, the solution to mathematical problems using pools of DNA molecules for data storage and parallel manipulations was implemented via human-directed reactions [1]. In particular, a computational program was encoded by an experimental protocol requiring human intervention. Subsequently, the feasibil- ity of computations employing DNA molecules to encode both data and instruction sets was investigated, and DNA hairpin formation was utilized in the solution to a combi- natorial problem instance for one-time only operation [2]. For successive operations, the use of restriction enzymes on double-stranded (ds) DNA molecules was proposed to implement a DNA-based automaton [3], in which a program was encoded by each reaction mixture involving a set of DNA molecules and enzymes. In Whiplash PCR (WPCR), successive operations are processed according to data and a computational program encoded within a single molecule [4]. Successive state transitions are implemented in parallel by the recursive, self-directed polymerase ex- tension of a mixture of single-stranded (ss) DNA molecules. In principle, WPCR is ⋆ to whom correspondence should be addressed. URL: http://www.apu.ac.jp/ ∼ jarose/.