Point-of-care COVID-19 testing: colorimetric diagnosis using rapid and ultra-sensitive ramified rolling circle amplification.

In this paper, we report a molecular diagnostic system-combining a colorimetric probe (RHthio-CuSO4) for pyrophosphate sensing and isothermal gene amplification (ramified rolling circle amplification)-that operates with high selectivity and sensitivity for clinical point-of-care diagnosis of SARS-CoV-2. During the polymerase phase of the DNA amplification process, pyrophosphate was released from the nucleotide triphosphate as a side product, which was then sensed by our RHthio-CuSO4 probe with a visible color change. This simple colorimetric diagnostic system allowed highly sensitive (1.13 copies/reaction) detection of clinical SARS-CoV-2 within 1 h, while also displaying high selectivity, as evidenced by its discrimination of two respiratory viral genomes (human rhino virus and respiratory syncytial virus) from that of SARS-CoV-2. All of the reactions in this system were performed at a single temperature, with positive identification being made by the naked eye, without requiring any instrumentation. The high sensitivity and selectivity, short detection time (1 h), simple treatment (one-pot reaction), isothermal amplification, and colorimetric detection together satisfy the requirements for clinical point-of-care detection of SARS-CoV-2. Therefore, we believe that this combination of a colorimetric probe and isothermal amplification will be useful for point-of-care testing to prevent the propagation of COVID-19.

Unnatural nucleotide-based rkDNA probe combined with graphene oxide for detection of alkaline phosphatase activity.

In this study we developed a fluorescent double-stranded DNA, incorporating an unnatural dUrk nucleotide, that we used as a probe for the detection of alkaline phosphatase (ALP) based on enzymatic cleavage of the non-fluorescent complementary strand. Primer extension performed using the unnatural nucleotide triphosphate dUrkTP and the natural deoxynucleotide triphosphates dATP, dCTP, and dGTP provided a simple fluorescent DNA strand that hybridized with the 5́-monophosphate non-fluorescent complementary strand. When applying the 5́-phosphate recognition and cleavage properties of lambda exonuclease (λ-exo), this probe could bind to graphene oxide (GO) and quench the fluorescence (in the absence of ALP) or not bind to GO and retain its fluorescence (in the presence of ALP). We obtained strongly fluorescent DNA strands through simple incorporation of multiple A sites in the complementary sequence, thereby increasing the number of dUrk residues during primer extension. This unnatural nucleotide-based rkDNA probing system exhibited high fluorescence differentiation for discriminating the status of ALP. This rkDNA-GO probing system appears to be a promising tool for monitoring the activity of disease-associated enzymes.

A fluorescent molecular rotor for the selective detection of the hybrid-conformation 22AG G-Quadruplex.

G-quadruplex (G4) DNA plays a vital role in myriad biological process and is linked to several human diseases, including Alzheimer's disease. Probing G4s with fluorescent probes can provide a better understanding their mechanisms of action and of their roles in Nature. In this study we developed a quinolinium-vinylaniline molecular rotor probe, featuring a diethylaminosalicylaldehyde unit that could discriminate the hybrid-22AG G4 sequence selectively amongst other G4 sequences. This probe underwent a significant red-shift upon binding to the target G4 (broad 575 nm â†’ sharp 630 nm) with enhanced fluorescence (up to 14-fold). We suspect that the vinylaniline unit of the molecular rotor, when bound to the hybrid-22 A G4, experienced restricted rotation, thereby undergoing enhanced intramolecular charge transfer. The presence of the diethylaminosalicylaldehyde moiety appeared to play a major role in the enhanced selectivity toward the 22AG G4.

Correction: Polymerase-mediated synthesis of p-vinylaniline-coupled fluorescent DNA for the sensing of nucleolin protein-c-myc G-quadruplex interactions.

Correction for 'Polymerase-mediated synthesis of p-vinylaniline-coupled fluorescent DNA for the sensing of nucleolin protein-c-myc G-quadruplex interactions' by Guralamatta Siddappa Ravi Kumara et al., Org. Biomol. Chem., 2021, DOI: 10.1039/D1OB00863C.

An oxidative cross-dehydrogenative-coupling reaction in water using molecular oxygen as the oxidant: vanadium catalyzed indolation of tetrahydroisoquinolines.

An aerobic oxidative cross-dehydrogenative coupling reaction between sp(3) C-H and sp(2) C-H bonds is developed by employing a vanadium catalyst (10 mol%) in an aqueous medium using molecular oxygen as the oxidant. This environmentally benign strategy exhibits larger substrate scope and shows high regioselectivity.