Label-free fluorescence strategy for methyltransferase activity assay based on poly-thymine copper nanoclusters engineered by terminal deoxynucleotidyl transferase.

The assay of detecting DNA methyltransferase activity has been identified as one of the central challenges in cancer diagnostics as DNA methylation is closely related to the diagnosis and treatment of tumors. In this study, a label-free fluorescence probe based on poly-thymine copper nanoclusters engineered by terminal deoxynucleotidyl transferase is proposed for methyltransferase activity assay. Taking advantage of the efficient polymerization extension reaction catalyzed by terminal deoxynucleotidyl transferase and the copper nanoclusters with large Stokes shift instead of labeling fluorescent dyes, the strategy exhibits a broader linear scope from 1 to 300 U mL-1 with a detection limit of 0.176 U mL-1. The economical method is specificity for M.SssI and also provides a convenient and high-throughput platform for screening the DNA methylation inhibitors, which displays a great potential for the practical applications of the drug development and clinical cancer diagnosis in the future.

Label-free and enzyme-free one-step rapid colorimetric detection of DNA methylation based on unmodified gold nanoparticles.

DNA methylation has been identified as one of the important causes of tumorigenesis, so it is important to develop some advanced methods for detecting and quantifying DNA methylation. In this study, a label-free and enzyme-free one-step rapid colorimetric detection of DNA methylation based on unmodified Au nanoparticles(Au NPs)has been proposed. This method can quickly, efficiently, economically and easily colorimetric detect methylated DNA only by the color change of unmodified Au NPs solution without the covalent modification of Au NPs in advance or complicated instruments for implementation with practical limitations or expensive biological enzymes or traditional organic dyes during the reaction. The strategy employed the difference in electrostatic attraction of single-stranded DNA and double-stranded DNA against salt-induced aggregation of Au NPs. The method has a DNA methylated detection limit of 8.47 nM and it is distinctly visible to detect methylated DNA with the naked eye as low as 20 nM. Furthermore, the strategy has an ability to detect methylated DNA in the presence of abundant unmethylated DNA with the detection limit of 0.13% and as low as 1% methylated DNA can be distinguished in heterogeneous samples with the naked eye. Also, the stratagem provides a convenient and rapid platform for methylated DNA detection of human serum samples in one step, which displays a huge potential for clinical diagnosis and treatment of oncological diseases.

Colorimetric determination of the activity of methyltransferase based on nicking enzyme amplification and the use of gold nanoparticles conjugated to graphene oxide.

A method is described for the colorimetric determination of the activity of CpG methyltransferase (M.SssI). It is based on (a) the crosslinking effect between dsDNA-modified gold nanoparticles (AuNPs) and graphene oxide (GO), and (b) an amplification reaction with the aid of a nicking enzyme. To avoid the aggregation of AuNPs (which would produce false signals), a hairpin DNA was connected to the AuNPs. Thus, the red color of the solution (measured at 530 nm) increases linearly with the activity of M.SssI from 0.2 to 60 U·mL-1, and the limit of detection is 67 U·mL-1. This is superior to some reported strategies. The method was successfully applied to analyze spiked serum samples. Conceivably, it represents a powerful tool for use in drug development and diagnosis. Graphical abstracts A method based on the conjugated cross-linking effect between dsDNA modified Au NPs and GO coupled with an amplification reaction of nicking enzyme has been developed for colorimetric detection of the activity of CpG methyltransferase (M.SssI).

Metal-Free Site-Specific Hydroxyalkylation of Imidazo[1,2- a]pyridines with Alcohols through Radical Reaction.

An effective approach to realize the direct hydroxyalkylation of imidazo[1,2- a]pyridines with alcohols promoted by di- tert-butyl peroxide was described without any metal catalyst. It is the first time that the dehydrogenative C(sp3)-C(sp2) coupling of imidazo[1,2- a]pyridines with alcohols occurred regioselectively at the C-5 position of imidazo[1,2- a]pyridines. Multisubstituted imidazopyridine derivatives were smoothly synthesized in moderate to good yields. Through a series of control experiments, a free-radical pathway was proposed to explain the experiment.

Copper-Mediated Oxidative Functionalization of C(sp3)-H Bonds with Isoquinolines: Two-Step Synthesis of 5-Oxaprotoberberinones.

A copper-mediated oxidative functionalization of C(sp3)-H bonds with isoquinolines via a radical process without ligands was achieved. The present system exhibits a novel pathway for the preparation of N-alkyl (benzyl) isoquinolin-1(2H)-ones in moderate to high yields. In addition, this procedure provides a simple method to afford 5-oxaprotoberberinones and their derivatives in two steps.