Figure 4.

Enhancing selectivity via competitive inhibition using an unstructured scavenger strand. (a) Without scavenger, HCR system H^U is not selective for cognate target X^A (forming Watson–Crick pair U·A) over 1-nt sequence variant X^G (forming nearly isoenergetic wobble pair U·G) (panel c: lanes 1 and 2). (b) Scavenger S^C is selective for X^G, restoring H^U selectivity for X^A via competitive inhibition (panel c: lanes 3 and 4). (c) Native PAGE poststained with SYBR Gold. Targets and scRNAs at 1 μM and scavenger at 2 μM. Reactions run for 1 h in 1× PKR at 37 °C. (d) Target sequences depicted 5′ to 3′ with nucleotide variant in orange. For cognate target X^A, the input domain of the cognate detecting scRNA is depicted with the location of the output domain indicated by an ellipsis. For off-target X^G, the scavenger S^C is depicted. See Supporting Information Tables S2 and S3 for scavenger and scRNA sequences; see Supporting Information Section S3 for additional data.