DNA-probes for the highly sensitive identification of single nucleotide polymorphism using single-molecule spectroscopy.
This article presents a new, highly sensitive method for the identification of single nucleotide polymorphisms (SNPs) in homogeneous solutions using fluorescently labeled hairpin-structured oligonucleotides (smart probes) and fluorescence single-molecule spectroscopy. While the hairpin probe is closed, fluorescence intensity is quenched due to close contact between the chromophore and several guanosine residues. Upon hybridization to the respective target SNP sequence, contact is lost and the fluorescence intensity increases significantly. High specificity is achieved by blocking sequences containing mismatch with unlabeled oligonucleotides. Time-resolved single-molecule fluorescence spectroscopy enables the detection of individual smart probes passing a small detection volume. This method leads to a subnanomolar sensitivity for this single nucleotide specific DNA assay technique.
Friedrich
Achim
A
Department of Functional Genome Analysis, German Cancer Research Center, Im Neuenheimer Feld 580, 69120 Heidelberg, Germany.
Hoheisel
Jörg D
JD
Marmé
Nicole
N
Knemeyer
Jens-Peter
JP
D001483:Base Sequence; D015342:DNA Probes / Q000737:chemistry*; D009693:Nucleic Acid Hybridization / Q000379:methods*; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods*; D013050:Spectrometry, Fluorescence / Q000379:methods*
D015342:DNA Probes