Oligonucleotide-based systems: DNA, microRNAs, DNA/RNA aptamers

Abstract

There are an increasing number of applications that have been developed for oligonucleotide-based biosensing systems in genetics and biomedicine. Oligonucleotide-based biosensors are those where the probe to capture the analyte is a strand of deoxyribonucleic acid (DNA), ribonucleic acid (RNA) or a synthetic analogue of naturally occurring nucleic acids. This review will shed light on various types of nucleic acids such as DNA and RNA (particularly microRNAs), their role and their application in biosensing. It will also cover DNA/RNA aptamers, which can be used as bioreceptors for a wide range of targets such as proteins, small molecules, bacteria and even cells. It will also highlight how the invention of synthetic oligonucleotides such as peptide nucleic acid (PNA) or locked nucleic acid (LNA) has pushed the limits of molecular biology and biosensor development to new perspectives. These technologies are very promising albeit still in need of development in order to bridge the gap between the laboratory-based status and the reality of biomedical applications.

Keywords: aptamers; biosensing; locked nucleic acids; microRNAs; oligonucleotides; peptide nucleic acids.

Figure 1.. Biogenesis and function of miRNAs

miRNAs are transcribed by RNA polymerase II into primary transcripts called pri-miRNAs which fold into a stem–loop structure. They are trimmed in the nucleus by the Microprocessor complex into pre-mRNAs. The latter are then exported with the aid of exportin 5. In the cytoplasm the enzyme Dicer further trims the pre-miRNAs into a mature miRNA duplex. The duplex is then complexed by Argonaute and other proteins forming mature microribonucleoprotein (miRNP) complexes (also referred to as miRNA-induced silencing complex (miRISC)) in which only the guide strand of the miRNA is retained. The guide strand is able to bind to mRNA targets in the cytoplasm. miRNAs are also found in processing bodies (P-bodies), cytoplasmic granules involved in mRNA turnover. (Reproduced from [2] by permission from Macmillan Publishers Ltd, copyright 2015.)

Figure 2.. Change in conformation from a single-stranded DNA aptamer to a quadruplex structure upon specific binding with thrombin

Figure 3.. Chemical structure of PNA and LNA

(A) Chemical model of a PNA molecule (sequence N-GTA-C) hybridized in antiparallel orientation with its complementary DNA (sequence 5′-TAC-3′). The dotted line indicates the hydrogen bonding between complementary nucleobases. (B) Chemical structure of an LNA monomer. (Adapted from [26] with permission from Springer Science and Business Media.)

Figure 4.. Schematic diagram showing the detection of miRNAs with nanoparticle amplified SPR detection

(i) specific hybridization of miRNA on to a complementary LNA array; (ii) addition of poly(A) tails to the surface-bound miRNAs using poly(A) polymerase enzyme; and (iii) hybridization of T₃₀-coated gold nanoparticles (Au NPs) to the poly(A) tails. (Reproduced with permission from [29], copyright 2006 American Chemical Society.)