Streptavidin-enhanced assay for sensitive and specific detection of single nucleotide polymorphism in TP53.
This paper reports an approach to detection of single nucleotide polymorphism based on special amplification assay and surface plasmon resonance biosensor technology. In this assay, a part of the target DNA is recognized by a probe (probe A) coupled with streptavidin-oligonucleotide (SON) complexes ex situ, and when the mixture is injected in the sensor, another part of the target DNA is recognized by a DNA probe (probe B) immobilized on the sensor surface. To achieve high sensitivity and specificity, the assay is optimized in terms of composition of SON complexes, probe design, and assay temperature. It is demonstrated that this approach provides high specificity (no response to targets containing single-mismatched bases) and sensitivity (improves sensor response to perfectly matched oligonucleotides by one order of magnitude compared to the direct detection method). The assay is applied to detection of a short synthetic analogue of TP53 containing a "hot spot"-single nucleotide mismatch frequently mutated in germ line cancer-at levels down to 40 pM.
Institute of Photonics and Electronics, Academy of Sciences of the Czech Republic, Chaberská, Prague, Czech Republic.
Hana
H
Sípová
Tomáš
T
Springer
Jiří
J
Homola
D015374:Biosensing Techniques / Q000379:methods; D004247:DNA / Q000235:genetics; D015342:DNA Probes / Q000235:genetics; D016158:Genes, p53 / Q000235:genetics*; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D009841:Oligonucleotides / Q000235:genetics; D020641:Polymorphism, Single Nucleotide / Q000235:genetics*; D012680:Sensitivity and Specificity; D019809:Streptavidin / Q000737:chemistry; D020349:Surface Plasmon Resonance / Q000295:instrumentation* / Q000379:methods*
D015342:DNA Probes; D009841:Oligonucleotides; D004247:DNA; D019809:Streptavidin