Split G-Quadruplexes Enhance Nanopore Signals for Simultaneous Identification of Multiple Nucleic Acids

Abstract

Assembly of DNA structures based on hybridization like split G-quadruplex (GQ) have great potential for the base-pair specific identification of nucleic acid targets. Herein, we combine multiple split G-quadruplex (GQ) assemblies on designed DNA nanostructures (carrier) with a solid-state nanopore sensing platform. The split GQ probes recognize various nucleic acid sequences in a parallel assay that is based on glass nanopore analysis of molecular structures. Specifically, we split a GQ into two asymmetric parts extended with sequences complementary to the target. The longer G-segment is in solution, and the shorter one is on a DNA carrier. If the target is present, the two separate GQ parts will be brought together to facilitate the split GQ formation and enhance the nanopore signal. We demonstrated detection of multiple target sequences from different viruses with low crosstalk. Given the programmability of this DNA based nanopore sensing platform, it is promising in biosensing.

Single Molecular Nanopores as a Label-Free Method for Homogeneous Conformation Investigation and Anti-Interference Molecular Analysis

“…For the product of 4H-A/T tail, the current blockage increases slightly, but the overall value does not exceed −300 pA (Figure S3e). According to the above and previous control experiments, 60,61 as shown in Figure 1e, the expected structures (S2 and S3) illustrated in Figure 1b form branched self-assembled structures (shortened as BAS), which are illustrated in Figure 2a,b.…”

“…For the product of 4H-A/T tail, the current blockage increases slightly, but the overall value does not exceed −300 pA (Figure S3e). According to the above and previous control experiments, , as shown in Figure e, the expected structures (S2 and S3) illustrated in Figure b may contribute those current blockages ≤400 pA, but should not have caused those super large current blockages ≥800 pA. Therefore, there is a rational prediction that under test conditions G-rich tail tag-induced S2–S2 and S3–S3 intermolecular assemblies may happen at different degrees to form branched self-assembled structures (shortened as BAS), which are illustrated in Figure a,b.…”

Experimental study on single biomolecule sensing using MoS2–graphene heterostructure nanopores

“…In recent decades, solid-state nanopores have emerged as a new tool with great potential in the detection of single biomolecules such as DNA, 1–9 RNA 10–12 and proteins. 13–15 In order to achieve single biomolecule sequencing and reveal more detailed information from the corresponding ionic blockage signals, the low spatial resolution and the ultra-fast translocation speed are two critical problems that remain to be solved.…”

Multiplexed Nanopore-Based Nucleic Acid Sensing and Bacterial Identification Using DNA Dumbbell Nanoswitches

“…by incorporating a logical arrangement of DNA dumbbell hairpins in a defined section on the carrier, allows the detection of multiple carriers using the same nanopore; therefore, the simultaneous detection of numerous target sequences can be achieved. However, in previous multiplexed nanopore sensing assays or data storage readout systems, ,− signals were identified by the presence or absence of a current peak at a specific position during carrier translocation. Thus, missing peaks are prone to lead to both false positive and negative results partly caused by velocity fluctuations during DNA translocation. , A more dynamic DNA nanostructure design that monitors changes in the structure should allow for multiplexed sensing with lower false positive rates.…”