Phenylethynylpyrene excimer forming hybridization probes for fluorescence SNP detection

Area of Science:

• Biochemistry
• Molecular Biology
• Fluorescence Spectroscopy

Background:

• Excimer formation in fluorescent dyes like pyrene offers unique biomolecule proximity sensing capabilities.
• Pyrene excimer fluorescence has been previously applied for homogeneous detection of single nucleotide polymorphisms (SNPs) in DNA.

Purpose of the Study:

• To evaluate the utility of 1-phenylethynylpyrene (1-1-PEPy) as a photostable, redshifted fluorescent probe for nucleic acid labeling.
• To demonstrate the application of 1-1-PEPy in excimer-forming DNA probes for detecting specific mutations in the 23S ribosomal RNA gene of Helicobacter pylori.

Main Methods:

• Synthesis and application of 1-phenylethynylpyrene (1-1-PEPy) derivatives for nucleic acid labeling.
• Utilizing excimer fluorescence of 1-1-PEPy for homogeneous detection of genetic variations.
• Targeting specific mutations (2144A/G, 2143A/G, 2143A/C) in the 23S rRNA gene of Helicobacter pylori.

Main Results:

• 1-1-PEPy forms excimers with emission maxima around 500-510 nm, suitable for nucleic acid labeling.
• Successful detection of specific 23S rRNA gene mutations associated with clarithromycin resistance in H. pylori strains using 1-1-PEPy DNA probes.
• Demonstrated generation of the phenylethynylpyrene pair from pseudonucleoside and modified nucleosides.

Conclusions:

• 1-Phenylethynylpyrene is a versatile and effective fluorescent label for developing excimer-forming DNA probes.
• This approach provides a sensitive method for detecting drug-resistant pathogens like Helicobacter pylori based on specific genetic markers.