Modular probes for enriching and detecting complex nucleic acid sequences

Abstract

Complex DNA sequences are difficult to detect and profile, but are important contributors to human health and disease. Existing hybridization probes lack the capability to selectively bind and enrich hypervariable, long or repetitive sequences. Here, we present a generalized strategy for constructing modular hybridization probes (M-Probes) that overcomes these challenges. We demonstrate that M-Probes can tolerate sequence variations of up to 7 nt at prescribed positions while maintaining single nucleotide sensitivity at other positions. M-Probes are also shown to be capable of sequence-selectively binding a continuous DNA sequence of more than 500 nt. Furthermore, we show that M-Probes can detect genes with triplet repeats exceeding a programmed threshold. As a demonstration of this technology, we have developed a hybrid capture method to determine the exact triplet repeat expansion number in the Huntington’s gene of genomic DNA using quantitative PCR.

Encoding quantized fluorescence states with fractal DNA frameworks

“…Structural DNA nanotechnology provides a powerful platform to construct self-assembled DNA nanostructures with nearatomic resolution and to programme the spatial organisation of molecules and nanoparticles with sub-10-nm resolution [22][23][24][25][26][27][28][29][30] . Specifically, the use of DNA frameworks as biocompatible scaffolds for programmable biomolecular assembly has been actively explored for various diagnostic, bioimaging and drug delivery applications 5,[31][32][33][34][35][36][37][38][39][40][41] . However, the regular, monomeric DNA frameworks do not satisfy the requirements for developing supermultiplex fluorescent amplifiers.…”

Simultaneous Detection of Different Zika Virus Lineages via Molecular Computation in a Point-of-Care Assay

“…The fact that multiple, different amplification reactions can be carried out in parallel, and yet the user need only attend to the final, fluorescent signal, highlights the power of the diagnostic, and presages the possibility of even more complex integration of molecular signals. For example, strand exchange nucleic acid computation has allowed (i) analysis of repetitive DNA targets of greater than 500 nucleotides in length with single nucleotide specificity, using “modular probes” composed of multiple short oligonucleotides that undergo competitive hybridization [56]; (ii) sensing and imaging of nucleic acids, enzymes, small molecules, proteins, and tumor cells via hairpin oligonucleotide probes and the hybridization chain reaction (HCR) [57]; and (iii) coupling nucleic acid detection to translational reporters via amplicon-mediated strand displacement reactions that couple to reporter protein production in a cell-free translation system [22].…”

Multiplex logic processing isothermal diagnostic assays for an evolving virus

“…The fact that multiple, different amplification reactions can be carried out in parallel, and yet the user need only attend to the final, fluorescent signal highlights the power of the diagnostic, and presages the possibility of even more complex integration of molecular signals. For example, strand exchange nucleic acid computation has allowed (i) analysis of repetitive DNA targets of greater than 500 nucleotides in length with single nucleotide specificity, using 'modular probes' composed of multiple short oligonucleotides that undergo competitive hybridization; 42 (ii) sensing and imaging of nucleic acids, enzymes, small molecules, proteins, and tumor cells via hairpin oligonucleotide probes and the hybridization chain reaction (HCR); 43 and (iii) coupling nucleic acid detection to translational reporters via amplicon-mediated strand displacement reactions that couple to reporter protein production in a cell-free translation system.…”