Detecting DNA at crime scenes using cascade toehold-mediated strand displacement

DNA sequencing is one of the main tools for identification of persons who were present at a crime scene. However, it is difficult to decide from which items within the crime scene samples should be collected. We propose to use cascade toehold-mediated strand displacement (CTMSD) to detect DNA at crime scenes. CTMSD consists of a sensing unit, composed of 3 short ssDNA molecules, and a fuel strand (FS), that enables recycling of the target molecule. The long strand (LS) of the sensing unit contains a sequence complementary to the target sequence (TS). The sensing strand (SS) of the sensor unit is released from the sensing unit in the presence of target DNA. Upon its release, the SS forms G-quadruplexes. In the presence of N-methyl mesoporphyrin IX (NMM), a commercially available porphyrin that is sensitive to G-quadruplexes (but not ssDNA or dsDNA), the released SS results in a strong fluorescent signal (with peak at 610 nm). We present how CTMSD can be adapted for detection of DNA at crime scenes.