5-Formylcytosine (5fC) is identified as one of the key players in active DNA demethylation and also as an epigenetic mark in mammals, thus representing a novel attractive target to chemical intervention. The current study represents an attempt to develop a reversible 5fC-targeted intervention tool. A supramolecular aldehyde reactive probe was therefore introduced for selective conversion of the 5fC to 5fC-AD nucleotide. Using various methods, we demonstrate that cucurbit[7]uril (CB7) selectively targets the 5fC-AD nucleotide in DNA, however, the binding of CB7 to 5fC-AD does not affect the hydrogen bonding properties of natural nucleobases in duplex DNA. Importantly, CB7-driven host–guest chemistry has been applied for reversible intervention of a variety of 5fC-targeted biochemical reactions, including restriction endonuclease digestion, DNA polymerase elongation, and polymerase chain reaction. On the basis of the current study, the macrocyclic CB7 creates obstructions that, through steric hindrance, prevent the enzyme from binding to the substrate, whereas the CB7/5fC-AD host–guest interactions can be reversed by treatment with adamantanamine. Moreover, fragment- and site-specific identification of 5fC modification in DNA has been accomplished without sequence restrictions. These findings thus show promising potential of host–guest chemistry for DNA/RNA epigenetics.

Natural nucleic acid bases can form Watson–Crick (WC) or Hoogsteen (HG) base pairs. Importantly, 8-oxo-2′-deoxyguanosine (8-oxo-dG) in DNA or 8-oxo-dG 5′-triphosphate (8-oxo-dGTP) favors a syn conformation because of the steric repulsion between O8 and O4′ of the deoxyribose ring. 8-oxo-dGTP can be incorporated into DNA opposite the templating adenine (A) using HG pairing as the dominant mechanism. Both RNA and DNA can be methylated at the N6 position of A to form N6-methyladenine (m6A). It has been found that certain viral infections may trigger an increase in the production of both 8-oxo-dGTP and m6A. The current study aims to systematically explore the effects of m6A methylation on HG base pairs and the consequent nucleotide incorporation. Our thermodynamic melting study shows that the m6A·8-oxo-dG is significantly less stable than the A·8-oxo-dG base pair in the paired region of a DNA duplex. Moreover, we have used pre-steady-state kinetics to examine the incorporation of 8-oxo-dGTP opposite m6A relative to A by a variety of reverse transcriptase (RT) enzymes and DNA polymerase (DNA pol) enzymes such as the human immunodeficiency virus type 1 (HIV-1) RT and human DNA pol β. The results demonstrate that all of these enzymes incorporate 8-oxo-dGTP less efficiently opposite m6A relative to A. Considering the steric bulk of the purine–purine pair between 8-oxo-dG and A, m6A methylation may affect the HG pairing to a great extent. Hence, it will be unfavorable to incorporate 8-oxo-dGTP into the growing strand opposite m6A. Moreover, the impeded incorporation of 8-oxo-dGTP opposite m6A has been extended to determine m6A at pre-defined positions in human rRNA. Our study may provide new insights into the roles of m6A in reducing the mutagenic potential of cellular 8-oxo-dGTP.

The epigenetic modification of nucleic acids represents one of the most significant areas of study in the field of nucleic acids because it makes gene regulation more complex and heredity more complicated, thus indicating its profound impact on aspects of heredity, growth, and diseases. The recent characterization of epigenetic modifications of DNA and RNA using chemical labelling strategies has promoted the discovery of these modifications, and the newly developed single-base or single-cell resolution mapping strategies have enabled large-scale epigenetic studies in eukaryotes. Due to these technological breakthroughs, several new epigenetic marks have been discovered that have greatly extended the scope and impact of epigenetic modifications in nucleic acids over the past few years. Because epigenetics is reversible and susceptible to environmental factors, it could potentially be a promising direction for clinical medicine research. In this review, we have comprehensively discussed how these epigenetic marks are involved in disease, including the pathogenesis, prevention, diagnosis and treatment of disease. These findings have revealed that the epigenetic modification of nucleic acids has considerable significance in various areas from methodology to clinical medicine and even in biomedical applications.

N 6-Methyladenine (m6A) is the most abundant internal modification on mammalian mRNA. Very recently, m6A has been reported as a potentially important ‘epigenetic’ mark in eukaryotes. Until now, site-specific detection of m6A is technically very challenging. Here, we first reveal that m6A significantly hinders DNA- and RNA-directed DNA synthesis. Systematic investigations of 5′-triphosphates of a variety of 5-substituted 2′-deoxyuridine analogs in primer extension have been performed. In the current study, a quantitative analysis of m6A in the RNA or DNA context has been achieved, using Bst DNA polymerase catalyzed primer extension. Molecular dynamics study predicted that m6A in template tends to enter into and be restrained in the MGR region of Bst DNA polymerase, reducing conformational flexibility of the DNA backbone. More importantly, a site-specific determination of m6A in human ribosomal RNA (rRNA) with high accuracy has been afforded. Through a cumulative analysis of methylation alterations, we first reveal that significantly cancer-related changes in human rRNA methylation were present in patients with hepatocellular carcinoma.

Here, we have provided novel insights into the effects of 8-oxodG substitutions on B–Z transitions of CpG dinucleotide DNAs. A combination of different techniques including a CD study, a PAGE analysis, DFT calculations and molecular modeling has been collectively used.

MicroRNA (miRNA) detection is of considerable significance in both disease diagnosis and in the study of miRNA function. The importance of miRNA itself is due to the complicated regulatory functions it plays in various life processes and its close relationship with some diseases. Traditional methods for miRNA detection do not meet the current demands, so various novel methods have been developed with a special focus on sensitivity and specificity. Herein, we summarize and discuss the newly developed miRNA detection methods.

G-triplex has recently been identified as a new secondary structure in G-rich sequences. However, its functions and biological roles remain largely unknown. This study first developed two kinds of Amplex Red oxidases, which were based on relatively new G-triplex structure and a common G-quadruplex one. A collection of DNA binding assays including circular dichroism (CD) spectroscopy, a CD melting assay, and a UV titration study were used to determine the G-triplex structure of G3 oligomer. The low intrinsic oxidative activity of hemin was significantly enhanced using G-triplex or G-quadruplex. Only one key guanine deletion from the G3 oligomer or G4 one could result in a much decreased Amplex Red oxidation activity. To the best of our knowledge, this is the first case reporting direct use of air as the oxidant for fluorescence generation based on DNAzyme strategies. Further mechanism studies demonstrated an involvement of on-site H2O2 generation from O2 and water and a following oxidation of Amplex Red to resorufin, causing a fluorescence enhancement. Furthermore, the newly developed oxidases have been effectively used in microRNA detection, using only one biotin-labeled probe and one small-molecule substrate. The conjugation of a target DNA to the G-triplex- or G-quadruplex-forming sequence enabled one to produce G-triplex or G-quadruplex by endonuclease in the presence of a slight amount of miRNA and amplify the signal of fluorescence from the oxidation of Amplex Red. Our findings of novel Amplex Red oxidases could potentially be used in a wide range of applications.

We have first demonstrated the distinctive effects of three newly reported epigenetic modifications, including 5hmC, 5fC, and 5caC, on B-Z transition of CpG dinucleotide DNAs. We have performed detailed assays and compared their effects. We further studied the regulation of B-Z transition of CpG dinucleotide dodecamers by alternating oxidation and alternating reduction.

This study is the first to investigate the interactions of hemin with a G-triplex DNA of T1 using different DNA binding assays. The low intrinsic peroxidatic activity of hemin could be significantly enhanced using T1. Furthermore, much decreased oxidation enhancement by T2 or T3 with one key guanine mutation was observed, and the observed peroxidatic activity of T1 should be directly due to the G-triplex complexed with hemin.

The epigenetic modification of nucleic acids represents one of the most significant areas of study in the field of nucleic acids because it makes gene regulation more complex and heredity more complicated, thus indicating its profound impact on aspects of heredity, growth, and diseases. The recent characterization of epigenetic modifications of DNA and RNA using chemical labelling strategies has promoted the discovery of these modifications, and the newly developed single-base or single-cell resolution mapping strategies have enabled large-scale epigenetic studies in eukaryotes. Due to these technological breakthroughs, several new epigenetic marks have been discovered that have greatly extended the scope and impact of epigenetic modifications in nucleic acids over the past few years. Because epigenetics is reversible and susceptible to environmental factors, it could potentially be a promising direction for clinical medicine research. In this review, we have comprehensively discussed how these epigenetic marks are involved in disease, including the pathogenesis, prevention, diagnosis and treatment of disease. These findings have revealed that the epigenetic modification of nucleic acids has considerable significance in various areas from methodology to clinical medicine and even in biomedical applications.

This study is the first to investigate the interactions of hemin with a G-triplex DNA of T1 using different DNA binding assays. The low intrinsic peroxidatic activity of hemin could be significantly enhanced using T1. Furthermore, much decreased oxidation enhancement by T2 or T3 with one key guanine mutation was observed, and the observed peroxidatic activity of T1 should be directly due to the G-triplex complexed with hemin.

N 6-Methyladenine (m6A) is the most abundant internal modification on mammalian mRNA. Very recently, m6A has been reported as a potentially important ‘epigenetic’ mark in eukaryotes. Until now, site-specific detection of m6A is technically very challenging. Here, we first reveal that m6A significantly hinders DNA- and RNA-directed DNA synthesis. Systematic investigations of 5′-triphosphates of a variety of 5-substituted 2′-deoxyuridine analogs in primer extension have been performed. In the current study, a quantitative analysis of m6A in the RNA or DNA context has been achieved, using Bst DNA polymerase catalyzed primer extension. Molecular dynamics study predicted that m6A in template tends to enter into and be restrained in the MGR region of Bst DNA polymerase, reducing conformational flexibility of the DNA backbone. More importantly, a site-specific determination of m6A in human ribosomal RNA (rRNA) with high accuracy has been afforded. Through a cumulative analysis of methylation alterations, we first reveal that significantly cancer-related changes in human rRNA methylation were present in patients with hepatocellular carcinoma.

MicroRNA (miRNA) detection is of considerable significance in both disease diagnosis and in the study of miRNA function. The importance of miRNA itself is due to the complicated regulatory functions it plays in various life processes and its close relationship with some diseases. Traditional methods for miRNA detection do not meet the current demands, so various novel methods have been developed with a special focus on sensitivity and specificity. Herein, we summarize and discuss the newly developed miRNA detection methods.

Here, we have provided novel insights into the effects of 8-oxodG substitutions on B–Z transitions of CpG dinucleotide DNAs. A combination of different techniques including a CD study, a PAGE analysis, DFT calculations and molecular modeling has been collectively used.