Binding-induced DNA assembly for detection of proteins and nucleic acids

• Author / Creator

  Lin,Yanwen

• The major research goal of my thesis is to design and develop the binding-induced assembly for the detection of specific proteins and nucleic acid sequences. My research builds on fundamentals of DNA chemistry and incorporates the recent advances in nanotechnology and signal amplification. I have studied the formation of binding-induced DNA three-way junctions (TWJs). By combining a TWJ with DNA strand displacement, I have developed a fluorescence turn-on assay and applied this assay to the detection of prostate specific antigen (PSA) and human α-thrombin. I used the binding-induced DNA TWJ to construct a molecular translator that converted the input target information into a pre-designed output DNA. The output DNA was hybridized to the oligonucleotides that functionalized on gold nanoparticles (AuNPs), each AuNP consisting of approximately 10^5 Au atoms. Detection of Au in the AuNPs using inductively coupled plasma mass spectrometry (ICP-MS) provided amplified and quantitative detection of the target molecules, such as human α-thrombin, microRNA 10b and microRNA 128. I further developed a binding-induced molecular amplifier (BIMA) technique, achieving improved sensitivity. AuNPs were used as the scaffold to conjugate multiple oligonucleotide sequences. Binding of a single target molecule to two affinity probes, one conjugated on the AuNPs and the other free in solution, induced the formation of a DNA structure with a recognizable nicking site. A nicking endonuclease enzyme cleaved off multiple DNA barcode molecules from each AuNP. The output DNA barcodes subsequently triggered the toehold-mediated strand displacement reactions that turned on fluorescence of molecular beacons. Detection of the amplified fluorescence signals enabled highly sensitive detection of the target molecules, including specific DNA sequences, PSA, platelet-derived growth factor (PDGF-BB), and the interaction between PDGF and its receptor β.The binding-induced DNA nanotechnologies described in this thesis have shown feasibility for the detection of several target proteins and nucleic acids. They have the promising potential for the detection of various biomolecules and applications to disease diagnostics.

• Subjects / Keywords

  • DNA three-way junction
  • gold nanoparticle
  • binding-induced molecular amplifier
  • binding-induced DNA assembly

• Graduation date

  Spring 2019

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-p4xp-w419

• License

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