Development and Applications of Fluorescent Oligonucleotides

Abstract

Fluorescent oligonucleotides (FONs) are used in a wide variety of areas such as molecular and mechanistics biological studies, molecular diagnostics, therapeutic development, biotechnology and nanotechnology. Ever since the post-genome era, there has been an ever-increasing demand for more rapid and accurate nucleic acid detection and quantification methods. Genetic information analyses require highly sensitive and specific detection of certain sequences, single nucleotide changes, specific structures and varied reactions in different formats in vitro, in living cells and, ultimately in animals and in human beings. Ideally, a unique event could be detected and quantified using the genomic information without amplification of the nucleic acids to be analyzed. Recent developments enabling detection at the single-molecule (SM) level have opened new perspectives for applications. This review reports on the development and applications of different families of FON probes. Specific insight is given to those leading to an important fluorescent signal change upon hybridization with their targets. Applications of FON probes include real time polymerase chain reaction (PCR) quantification, detection of single-nucleotide polymorphisms (SNP), fluorescence in situ hybridization (FISH) including detection of specific messenger RNAs in living cells, analysis of gene expression, analysis of nucleic acid structures and reactions, and in nanotechnology, the assessment of molecular machine motion and functioning.

Synthesis and spectral characterization of environmentally responsive fluorescent deoxycytidine analogs

“…[1][2][3][4][5] Naturally occurring nucleobases are virtually non-fluorescent under ambient conditions 6 and thus require chemical modification to be endowed with desirable fluorescence properties. Such fluorescent properties may come from attachment of a pendent fluorophore (e.g.…”

Recent developments in oligonucleotide conjugation

“…Use of complementary ONs labelled with different fluorescent labels, emitting at different wavelengths, enables the concomitant detection of several genes in the same experiment. 18 Molecular beacons (MBs) are widely used in chemistry, biology, biotechnology, and medical sciences for biomolecular recognition, due to their ease of synthesis, unique functionality and molecular specificity. 20 MBs act like switches; they are non-fluorescent in the free form (fluorescence ''off'') but exhibit strong fluorescence (fluorescence ''on'') when bound to target sequences.…”

“…MBs have also been used for investigating the tertiary DNA structures such as triplex and G-quadruplex DNA. 18 Furthermore, the high selectivity of MBs enables the simple, rapid and sensitive detection of single-nucleotide polymorphisms (SNPs). 18 MBs have been used for detecting protein-DNA interactions (single-stranded DNA binding protein or SBB from E. coli), monitoring the activity of enzymes (E. coli DNA ligase), and detecting small molecules (ATP).…”

Rapid genotyping using pyrene−perylene locked nucleic acid complexes

“…5 Förster resonance energy transfer (FRET) is one of the major fluorescence effects applied in nucleic acid biosensing which has been studied thoroughly. [6][7][8][9][10][11][12][13][14][15][16][17][18][19] Being a physical process whereby the excited state energy of a donor molecule can be transferred to a neighboring acceptor fluorophore in the ground state, FRET can take place whenever the two fluorophores with sufficient spectral overlap between the donor fluorescence and acceptor absorbance are in close proximity (distance typically below 7 nm), and importantly, provided favorable donor-acceptor dipole-dipole orientation. FRET can be monitored quantitatively and in real time (e.g., real-time PCR), [20][21][22][23][24][25][26] with additional possibilities for…”