Fluorogenic minor groove binder (MGB) probes containing an MGB-quencher at the 5'-end and a fluorophore at the 3'-end have been recently reported. These probes fluoresce on hybridization to the complementary targets. The 5'-MGB-quencher group prevents 5' - nuclease digestion by Taq polymerase during homogeneous amplification. The 5'-MGB-quencher-oligonucleotide-fluors can be used as probes in general nucleic acid and SNP detection assays. We propose to use a new software prediction program to develop MGB Eclipse probe assays to detect Clostridium botulinum, Bacillus anthracis and Yersinia pestis. The designed probes and primers will be optimized with use of three modified bases to improve probe performance of AT- and GC-rich sequences. In the Phase 1 effort, we will design and optimize the primers and probes and demonstrate function of the MGB Eclipse probe assays on a set of known target samples. The specificity of the probes will be evaluated against panels of closely related non-pathogenic organisms. In Phase II, we will expand the number of Category A organisms and improve throughput by multiplexing. In addition, commercial prototype MGB Eclipse(TM) probe assays will be developed for each of these organisms.
Thesaurus Terms: biohazard detection, biomarker, fluorescent dye /probe, nucleic acid probe, technology /technique development Bacillus anthracis, Clostridium botulinum, DNA primer, Yersinia pestis, nucleic acid hybridization, single nucleotide polymorphism biotechnology, bioterrorism /chemical warfare, polymerase chain reaction
