Part:BBa_K2965002

H2 (Hybridization chain reaction)

H2 is a single strand DNA, which participates in part of the hybridization chain reaction together with H1(BBa_K2965001) when triggered by S2 (BBa_K2965003). We use it to realize signal amplification in hybridization chain reaction.

Usage and Biology

Hybridization chain reaction (HCR) is a simple and efficient isothermal amplification process proposed by Dirks and Pierce in 2004. In a typical HCR, an initiator triggers a cascade of hybridization events between two species of DNA hairpins, leading to the formation of a nicked double helix with tens to hundreds of repeated units until the hairpins are exhausted. Given the advantages of its enzyme-free nature, efficient isothermal amplification, ultrahigh sensitivity and structural flexibility, HCR has emerged as a powerful molecular tool with versatile applications in biosensing, bioimaging, and biomedicine. Biosensing, signal transduction, or transforming input molecules into readable signal outputs, can readily be achieved via incorporation of HCR products with functional moieties such as fluorophores, nanoparticles (NPs), and electrochemical reagents.

Fig1. Hybridization chain reaction.An initiator (S2) triggers a cascade of hybridization events between two species of DNA hairpins (H1, H2), leading to the formation of a nicked double helix with tens to hundreds of repeated units until the hairpins are exhausted.

Characterization

Result

Characterization of the HCR of pure hairpin DNA strands is as shown in Fig2. Without the initiator (S2), the hybridization chain reaction cannot occur. When initiator (S2) exists, the hybridization chain reaction occurs, and the length nicked double helix varies with the concentration of initiator (S2).

Fig2. Characterization of the HCR of pure hairpin DNA strands by 8% native PAGE.(1)H1; (2)H2; (3)H1+H2; (4−9) H1+H2+S2; (10)DNA ladder (DL2000 plus). The concentration of H1 and H2 in lane 1, 2, 4 was 1μM, while in lane 5−9 was 2.5 μM. The concentrations of S2 was 1μM in lane 3, while 200 nM, 100 nM, 20 nM, 10 nM, 1 nM from lane 5 to 9, respectively.

Protocol

Probe DNA, H1 and H2 (10μM) were prepared in 1*NEBuffer 1 and heated at 90℃ for 10min separately, then they were allowed air cooling for 1 hour to form hairpin structure. In the hybridization chain reaction system, add 10*NEBuffer 1 to final concentration of 1*.

Reference

[1] Bi S, Yue S and Zhang S. Hybridization chain reaction: A versatile molecular tool for biosensing, bioimaging, and biomedicine. Chem Soc Rev. 2017; 46：

[2] Lu S, Hu T, Wang S, Sun J and Yang X. Ultra-Sensitive Colorimetric Assay System Based on the Hybridization Chain Reaction-Triggered Enzyme Cascade Amplification. Acs Appl Mater Inter. 2017; 9：167-175.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]