The Nanopore-Tweezing-Based, Targeted Detection of Nucleobases on Short Functionalized Peptide Nucleic Acid Sequences

Abstract

The implication of nanopores as versatile components in dedicated biosensors, nanoreactors, or miniaturized sequencers has considerably advanced single-molecule investigative science in a wide range of disciplines, ranging from molecular medicine and nanoscale chemistry to biophysics and ecology. Here, we employed the nanopore tweezing technique to capture amino acid-functionalized peptide nucleic acids (PNAs) with α-hemolysin-based nanopores and correlated the ensuing stochastic fluctuations of the ionic current through the nanopore with the composition and order of bases in the PNAs primary structure. We demonstrated that while the system enables the detection of distinct bases on homopolymeric PNA or triplet bases on heteropolymeric strands, it also reveals rich insights into the conformational dynamics of the entrapped PNA within the nanopore, relevant for perfecting the recognition capability of single-molecule sequencing.

Probing the Hepatitis B Virus E-Antigen with a Nanopore Sensor Based on Collisional Events Analysis

“…Measurements were carried out as previously reported [ 67 ]. The recording cell consisted of two chambers of 1 mL volume termed cis (grounded) and trans , separated by a 25 μm-thick Teflon film (Goodfellow, Malvern, MA, USA) containing an aperture of ~120 μm in diameter.…”

Artificial genetic polymers against human pathologies

“…However, to reach these goals, a lot of optimization steps are required. Applications of AGPs will be further improved through the use of special dedicated software [ 174 ], detection of PNAs using nanopore platforms [ 175 ], and with rational improvements in reducing side effects such as hepatotoxicity [ 176 ]. Machine learning algorithms partially help alleviate this problem by predicting possible side effects of experimental drugs, however, a lot of work needs to be done in this direction [ 177 – 179 ]…”

A Nanopore Sensor for Multiplexed Detection of Short Polynucleotides Based on Length-Variable, Poly-Arginine-Conjugated Peptide Nucleic Acids

“…As reviewed recently, at the very core of the nanopore-based, real-time, and specific detection of target polynucleotide species is the possibility to discern the blockade signature accompanying the following amplification-free, hybridization-driven duplex formation with a probe polynucleotide in a heterogeneous mix of polynucleotides. This creates an extremely effective opportunity to identifying specific single-stranded DNA (ssDNA) strands with nanopores presenting multiple base recognition sites spanning nanometer distances (e.g., the α-HL protein nanopore), circumventing the spatial resolution required for the actual sequencing, , by eliminating the effect of spatial uncertainty when the ssDNA strand probes the nanopore. By employing either conventional ssDNAs or synthetic peptide-nucleic acid (PNA) fragments containing an uncharged N -(2-aminoethyl)-glycine-based pseudopeptide backbone and capable of forming Watson–Crick complementary duplexes with biological DNA, the above-mentioned strategy was successful in detecting and probing at the single-molecule level the kinetics of DNA/DNA or DNA/PNA duplex formation ,− or miRNA-selective detection. ,, …”