Fig. 4

The conditional and structural optimization of the HCR efficiency of DSAP. a The influence of reaction temperature on the HCR efficiency of the DSAP, including 4, 25, and 37 °C. b The influence of Mg²⁺ concentrations in the reaction system on the HCR efficiency of the DSAP, including 2 mM Mg²⁺, 10 mM Mg^2+, and 50 mM Mg²⁺. c The influence of sticky end length on HCR efficiency of DSAP, including 13, 17, and 21 nt. The error bars in (a–c) are SD according to three repetitive experiments. d The AGE showing the initiator, DSAP, and the reaction products between initiators and DSAP of different ratios. The red bands are generated by the Cy5 channel, and the green bands are generated by the GelRed channel. As the Figure shows, the initiator of lower concentration can trigger larger DNA polymers, but the initiator of higher concentration can trigger smaller DNA polymers. e The morphology and particle sizes of the DSAP and HCR reaction products between DSAP and initiator at the molar ratios of 4:1 were determined by AFM imaging and particle size analysis. The mean particle size is decided by three parallel measurements. f The breathing sites can trigger self-hybridization of H1-SE and H2-SE. The base pair mismatches in the breathing sites of H2-SE can improve the metastability of DSAP. The visualization of the DSAP with no mismatch, one mismatch, and two mismatches was displayed. The AGE and fluorescence intensity measurements were also shown for the corresponding group. The error bars in (a–f) are determined by the SD of the MFI from at least five parallel experiments. All the tested samples were technical replicates. The “I” represents the initiator. The **** represented p < 0.0001, ** represented p < 0.01. Scale bars, 10 nm