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Nucleic acid modifications play essential roles in diverse biological processes, ranging from gene expression regulation to stress response. While traditional research focused on common modifications like methylation, recent discoveries are unveiling a wide range of rare modifications with potentially crucial functions. However, accurately detecting and mapping these modifications pose significant challenges due to their low abundance and diverse chemical properties. This article summarizes the recent discoveries of rare DNA and RNA modifications across various organisms, highlighting their potential biological significance. Furthermore, it critically evaluates the limitations of current mapping techniques, including potential sources of false positives and negatives. Finally, the article discusses emerging strategies for overcoming these challenges and future opportunities in the field of rare nucleic acid modification detection. [ABSTRACT FROM AUTHOR]

N6-methyladenosine (m6A) has been increasingly recognized as a new and important regulator of gene expression. To date, transcriptome-wide m6A detection primarily relies on well-established methods using next-generation sequencing (NGS) platform. However, direct RNA sequencing (DRS) using the Oxford Nanopore Technologies (ONT) platform has recently emerged as a promising alternative method to study m6A. While multiple computational tools are being developed to facilitate the direct detection of nucleotide modifications, little is known about the capabilities and limitations of these tools. Here, we systematically compare ten tools used for mapping m6A from ONT DRS data. We find that most tools present a trade-off between precision and recall, and integrating results from multiple tools greatly improve performance. Using a negative control could improve precision by subtracting certain intrinsic bias. We also observed variation in detection capabilities and quantitative information among motifs, and identified sequencing depth and m6A stoichiometry as potential factors affecting performance. Our study provides insight into the computational tools currently used for mapping m6A based on ONT DRS data and highlights the potential for further improving these tools, which may serve as the basis for future research. Direct RNA sequencing using nanopore platform can be used to detect N6-methyladenosine (m6A) modifications on mRNAs. Here the authors systematically compare tools used for m6A detection from nanopore direct sequencing. [ABSTRACT FROM AUTHOR]

N6-deoxyadenosine methylation (6mA) is the most widespread type of DNA modification in prokaryotes and is also abundantly distributed in some unicellular eukaryotes. However, 6mA levels are remarkably low in mammals. The lack of a precise and comprehensive mapping method has hindered more advanced investigations of 6mA. Here, we report a new method MM-seq (modification-induced mismatch sequencing) for genome-wide 6mA mapping based on a novel detection principle. We found that modified DNA bases are prone to form a local open region that allows capture by antibody, for example, via a DNA breathing or base-flipping mechanism. Specified endonuclease or exonuclease can recognize the antibody-stabilized mismatch-like structure and mark the exact modified sites for sequencing readout. Using this method, we examined the genomic positions of 6mA in bacteria (E. coli), green algae (C. reinhardtii), and mammalian cells (HEK239T, Huh7, and HeLa cells). In contrast to bacteria and green algae, human cells possess a very limited number of 6mA sites which are sporadically distributed across the genome of different cell types. After knocking out the RNA m6A methyltransferase METTL3 in mouse ES cells, 6mA becomes mostly diminished. Our results imply that rare 6mA in the mammalian genome is introduced by RNA m6A machinery via a non-targeted mechanism. [ABSTRACT FROM AUTHOR]

Stealth liposomes have been broadly investigated as drug delivery or diagnostic agent. However, the materials that possess the ability of stealth, such as DSPE-PEG and Chol-PEG, are either costly or synthetic complex. In this research with different molecular weights (2000-6000 g/mol) of methoxy poly(ethylene glycol)(MePEG), a series of MePEG-bile (MePB) conjugates were generated by an economical and simple method and confirmed by FTIR and 1H-NMR spectrum. The properties in aqueous solution were studied, including viscosity and surface activity, over a wide concentration range. To elucidate the application of MePB in liposomes (MePBL), conventional liposomes (CL) were prepared, and the influence of the grafting density and the chain length of MePB in liposomes were investigated. The ability of long circulation of MePBL was evaluated by intravenous injection administration in rats. Results indicated that all the liposomes prepared, with or without MePB composition, were similar in micrograph, and the contents of MePB in MePBL were more important than the chain length of MePB for a long circulation in vivo. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011 [ABSTRACT FROM AUTHOR]

Purpose: The objective of this study was to achieve a sustained and targeted delivery of liposome to the liver, by modifying the phospholipid [phosphatidylcholine (PC)/cholesterol (10 : 1) liposomes with a novel polymer bile salts-(polyethylene glycol)2000-bile salts (BP2B). Methods: First, we generated a novel BP2B by N,N′-dicyclohexylcarbodiimide/4‐dimethylaminopyridine esterification method and confirmed by Fourier transform infraredand H‐NMR spectra. Second, we prepared the BP2B-modified liposomes (BP2BL) that included BP2B, and the effect of the weight ratios of BP2B/PC on entrapment efficiency was investigated and BP2B/PC = 3% (w/w) was determined as the optimum ratio for the 4,4′-dimethoxy-5,6,5′,6′-bi (methylenedioxy)-2,2′-bicarbomethoxybiphenyl liposomes. And then, the ability of the liver target of BP2BL was studied by calculating the targeted parameters. Results and Discussion: All the results revealed that the introduction of polyoxyethylene chains could control interactions of bile salt moieties on liposome surfaces with the receptor compared with traditional liposomes (CL), marking BP2BL as a suitable carrier for hepatic parenchymal cell-specific and sustained targeting. It was suggested that liposomes containing such novel BP2B have great potential as drug delivery carriers for the liver-selective targeting that has targeted and sustained drug delivery. [ABSTRACT FROM AUTHOR]

The past few years have witnessed rapid progress in the field of RNA modifications. As the most prevailing modification on eukaryotic mRNA, m 6 A is characterized to play a vital role in various cellular activities. However, limitations of the detection method impede functional studies of m 6 A. Here we introduce m 6 A-REF-seq, a powerful and straightforward method to identify m 6 A at single-nucleotide resolution. m 6 A-REF-seq relies on the recognition of RNA endonuclease MazF towards m 6 A at the ACA motif, providing an orthogonal method independent of the m 6 A antibody being adopted by most of current methods. We describe a detailed protocol to perform m 6 A-REF-seq, including NGS library construction and sequencing data analysis. In particular, we describe an optimized assay to validate individual m 6 A sites identified by m 6 A-REF-seq, which can also be applied to detect any candidate m 6 A sites., (Copyright © 2021 Elsevier Inc. All rights reserved.)

The aim of this study was to develop a sustained release converse thermosensitive hydrogel for intra-articular injection using chitosan-glycerol-borax as matrix, its physical properties and biocompatibility were investigated. Taking gelation time and gelation condition as index, the influence of concentration of chitosan, ratio of chitosan to glycerol, pH on physical properties of hydrogel were investigated. And then the in vitro drug release, rheological properties and biocompatibility were studied. The thermosensitive hydrogel flows easily at room temperature and turns to gelation at body temperature, which can certainly prolong the release of drug and has good biocompatibility.

To explore the mechanism of the absorption enhancement of borneol, the effect of borneol on the intestinal absorption and the pharmacokinetics of tetramethylpyrazine phosphate after oral administration were investigated. In situ intestinal recirculation was performed to study the effect of various concentrations of borneol on the absorption of tetramethylpyrazine phosphate at duodenum, jejunum, ileum and colon. After oral administration of tetramethylpyrazine phosphate, the mixture of tetramethylpyrazine phosphate and borneol and the mixture of tetramethylpyrazine phosphate and verapamil in rats, the concentrations of tetramethylpyrazine phosphate in plasma were determined by RP-HPLC at predesigned time. The pharmacokinetic parameters were compared based on the results of the three animal experiments, and analyzed with software program 3p97. The result showed that tetramethylpyrazine phosphate could be absorbed at all of the four intestinal segments with increasing absorption amount per unit as follows: colon > duodenum > jejunum > ileum, but without saturation, which demonstrated that tetramethylpyrazine phosphate was absorbed via simple diffusion. Borneol could enhance the intestinal absorption of tetramethylpyrazine phosphate, however, not in proportion. There was no obvious difference between the test group and the control group when 10 microg x mL(-1) borneol was added (P > 0.05), while when the concentration comes to 25 microg x mL(-1) and 50 microg x mL(-1), significant differences were observed (P < 0.05). Borneol and verapamil did enhance the bioavailability of tetramethylpyrazine phosphate after oral administration in rats. The enhancing effect of borneol showed only when the concentration came to a certain level but with no specific sites existed in the intestine. One of the mechanisms of borneol on the enhancing effect on absorption of tetramethylpyrazine phosphate might be the inhibition of the metabolism of CYP 3A and exocytosis of P-gp.

Aim: To prepare verapamil hydrochloride controlled-onset extended-release pellets (VH-COERP) and study its release behavior in vitro. To compare the pharmacokinetic characteristics and bioavailability in six Beagle dogs after oral administration of VH-COERP and verapamil hydrochloride delayed-release pellets (VH-DRP) as reference., Methods: The core of VH-COERP were prepared in the fluidized bed (mini-glatt) by spraying water solution containing drugs onto sucrose-starch pellets with hydroroxy propyl methyl cellulose (HPMC) as the inner coating swelling layer and ethylcellulouse aqueous dispersion as the outer coating controlled layer. Through modifying the coating level of inner and outer layer, the VH-COERP with the optimized cumulative release profile was obtained. The concentration of VH in plasma of six dogs and its pharmacokinetic behaviors after oral administration of VH-COERP and VH-DRP at different times were studied by RP-HPLC. The pharmacokinetic parameters were computed by software program 3P97., Results: The lag time, the release behavior and the amount of VH from VH-COERP within 24 hours were not influenced by the pH of dissolution medium and post-process, but obviously influenced by the different kinds of added material in swelling layer and the coating level of the inner swelling layer and the outer controlled layer. In vitro the lag time of release profile of VH from VH-COERP was 5 h and then VH was extended release from VH-COERP in the following time. Compared with the VH-DRP, VH-COERP in vivo has an obviously lag time (4 h) , Tmax was also delayed (8 h) and the relative bioavailability was (94.56 +/- 7.64)%., Conclusion: The release profile of VH from VH-COERP was shown to be extended-release after an conspicuous lag time in vitro and in vivo. So the drug can be taken by the patient before bed time and begin to work at the morning.