Specific detection of bacterial pathogens using oligonucleotide microarrays generated from hydrolysis PCR probe sequences.
A move away from high-density screening array formats and the implementation of probes specifically identifying targets for application in low-density hybridization capture analyses is growing in importance. Some of the highest specificity for bioassays that encompasses use of hybridization has enlisted hydrolysis probes in real-time PCR. It is demonstrated that by employing 5'-end-tethering to glass of hydrolysis PCR probe sequences that these can be employed as capture probe sequences. The probes retain hybridization binding specificity to their PCR amplicons and specificity of hybridization was achieved across all probes at equivalent stringency. This suggests that probes identified for use in hydrolysis PCR using one set of temperature cycling parameter can also be applied to a microarray, where again the probes all exhibit similar specificity of interact at analogous hybridization conditions. The procedure permits the exact same PCR amplicon sequences to be used in both quantitative PCR and qualitative microarray assay formats.
Day
Philip
P
Manchester Interdisciplinary Centre, University of Manchester, UK.
D001419:Bacteria / Q000235:genetics / Q000472:pathogenicity*; D001424:Bacterial Infections / Q000175:diagnosis* / Q000235:genetics / Q000382:microbiology; D004269:DNA, Bacterial / Q000032:analysis; D006868:Hydrolysis; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D015345:Oligonucleotide Probes / Q000138:chemical synthesis / Q000592:standards*; D016133:Polymerase Chain Reaction / Q000379:methods*
D004269:DNA, Bacterial; D015345:Oligonucleotide Probes