Extreme DNA resolution: Researchers slow down and scan multiple times individual DNA molecules

(Nanowerk News) Aleksandra Radenovic, leader of the Nanoscale Biology Laboratory in the Engineering School, has dedicated years to enhancing nanopore technology, wherein a molecule such as DNA is guided through a minuscule aperture in a membrane to assess an ionic flow. Through analyzing the impact of each nucleotide on this flow as it traverses, scientists can decipher the sequence of DNA's nucleotides that encode genetic information.
The findings have been published in Nature Nanotechnology ("Spatially multiplexed single-molecule translocations through a nanopore at controlled speeds").
Presently, the movement of molecules through a nanopore and the temporal aspects of their analysis are subject to stochastic physical forces, while the swift motion of molecules poses difficulties in attaining precise analytical outcomes. Radenovic has previously tackled these challenges by employing optical tweezers and viscous fluids. However, through a collaboration with Georg Fantner and his team at EPFL's Laboratory for Bio- and Nano-Instrumentation, she has achieved the breakthrough she has long sought. The outcomes of this collaboration have the potential to extend well beyond DNA research.
"The amalgamation of nanopore sensitivity and scanning ion conductance microscopy (SICM) precision has enabled us to selectively target particular molecules and locations while regulating their movement velocity," Radenovic explains. "This remarkable level of control has the potential to address a significant void in the field." The team accomplished this feat by repurposing a cutting-edge scanning ion conductance microscope, a recent innovation from the Lab for Bio- and Nano-Instrumentation.

Improving sensing precision by two orders of magnitude

The fortuitous collaboration between the two laboratories was initiated by PhD student Samuel Leitão, whose research centers around scanning ion conductance microscopy (SICM). SICM involves leveraging fluctuations in the ionic current within a probing tip to generate detailed 3D imaging data. During his doctoral studies, Leitão focused on advancing and implementing SICM technology for imaging nanoscale cellular structures, utilizing a glass nanopore as the probing tool. In this recent endeavor, the team harnessed the precision of a SICM probe to direct the movement of molecules through a nanopore, instead of relying on random diffusion.
Referred to as scanning ion conductance spectroscopy (SICS), this breakthrough technique decelerates the passage of molecules through the nanopore, enabling the acquisition of numerous sequential readings from a single molecule, as well as from distinct positions along the molecule. By exerting control over the transit speed and accumulating multiple readings of the same molecule, the signal-to-noise ratio has witnessed a remarkable increase of two orders of magnitude when compared to conventional methods.
“What's particularly exciting is that this increased detection capability with SICS may be transferable to other solid-state and biological nanopore methods, which could significantly improve diagnostic and sequencing applications,” Leitão says.
Fantner succinctly captures the rationale behind the approach using an automotive analogy: "Imagine yourself observing cars passing by while you stand in front of a window. It becomes significantly easier to read their license plate numbers if the cars slow down and drive past repeatedly," he explains. "In a similar manner, we now have the ability to choose between measuring 1,000 different molecules once or obtaining 1,000 consecutive measurements of the same molecule. This represents a genuine paradigm shift in the field."
The high precision and adaptability of this approach open up possibilities for its application to molecules beyond DNA, including peptides, which are essential protein building blocks. This development holds the potential to propel advancements in proteomics, as well as biomedical and clinical research. By extending the scope of application, the research can contribute to a broader understanding of molecular structures and functions, with implications for various scientific disciplines and practical applications in healthcare.
Radenovic acknowledges that sequencing peptides has presented a notable challenge owing to the intricate nature of their "license plates," which consist of 20 distinct characters (amino acids) compared to the four nucleotides found in DNA. In light of this, she expresses her enthusiasm regarding the potential of this newfound control, as it may pave the way for a more streamlined approach to peptide sequencing. This prospect holds great promise in expanding our understanding of peptides and their roles, offering exciting opportunities for advancements in the field of proteomics.
Source: École Polytechnique Fédérale de Lausanne (EPFL) (Note: Content may be edited for style and length)
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