Fluorescence Detection of Specific RNA Sequences Using 2′-Pyrene-Modified Oligoribonucleotides

Fluorescent oligonucleotide probe based on G-quadruplex scaffold for signal-on ultrasensitive protein assay

“…from the removal of single-stranded segment of LFGQ probe/ aptamer hybrid. There are essentially three reasons for the observation: 1) although there are G:T mismatches, the SP1/aptamer hybrid should exhibit a relatively strong cleavage resistance because the G:T mismatch has been shown to cause only a relative minimum of structural disturbance within the DNA duplex [28,29]; 2) the S1 nuclease under chosen conditions could degrade single-stranded oligonucleotides while avoiding potential double-stranded DNA cleavage according to the previous work optimizing the nuclease concentration [22]; 3) the nucleobases serve as an efficient quencher for pyrene fluorescence, and the pyrene-labeled DNA/complementary sequence (including RNA sequence) hybrids display very weak monomer fluorescence [17,19]. The experimental results demonstrate, on the basis of competitive binding for the aptamer sequences, that LFGQ probe could be used as signaling probe for target species detection.…”

“…Utilizing the change in the local environment of pyrenes originating from the labeled DNA cleavage by S1 nuclease, the described interrogating strategy not only circumvents the intrinsic limitation for an efficient implementation associated with the quenching of the pyrene monomer excited singlet state by the flanking nucleobases in both noncovalent/covalent hybrids and single-stranded oligonucleotides [19,43] but also displays obvious advantages of rapidity, cost effectiveness, high sensitivity, high-throughput capability and so on. The novel conformation-switching signaling element capable of transducing efficiently the binding reaction into detectable physical signal could be employed to construct a wide range of biosensing schemes for the analysis of various target species via being mediated by a target-binding complementary aptamer; 2) the CFGQ probe designed in our work would open the way for the design and application of spatially sensitive signaling probe based on the singlepyrene excimer emission.…”

“…aptamer) undergoing a significant target-induced conformational change without obvious loss of binding activity, and the pyrenes attached to oligonucleotides often exhibit moderate or poor fluorescence due to quenching nucleobases [19].…”

REACTION OF N3-BENZOYL-3′,5′-O-(DI-TERT-BUTYLSILANEDIYL)URIDINE WITH HINDERED ELECTROPHILES: INTERMOLECULAR N3To 2′-OPROTECTING GROUP TRANSFER

“…However, published alkylations have been achieved only with relatively reactive electrophiles: for example, methyl iodide has been used frequently to methylate the ribosyl 2 0 -OH in reaction with silver oxide, [12][13][14] sodium hydride [15] or 2-tert-butylimino-2-diethylamino-1,3-dimethyl-perhydro-1,3,2-diazaphosphorine (BEMP). [16] Reactive benzyl and allyl halide derivatives [17][18][19][20] often have been incorporated into oligonucleotides via 2 0 -O-alkylation as antisense groups or fluorescent probes. [18] Protecting group agents such as alkoxymethyl halides [21] and alkyl bromoacetates [22][23][24] also have worked well, owing to their high reactivities.…”

“…[16] Reactive benzyl and allyl halide derivatives [17][18][19][20] often have been incorporated into oligonucleotides via 2 0 -O-alkylation as antisense groups or fluorescent probes. [18] Protecting group agents such as alkoxymethyl halides [21] and alkyl bromoacetates [22][23][24] also have worked well, owing to their high reactivities. Treating 2,2 0 -anhydrouridine with boron alkoxides has provided an alternative access to 2 0 -O-alkyluridine with less reactive electrophiles, [25,26] but high temperature ( > 100 C) required in this reaction 724 ZHU ET AL.…”