Development of high sensitive and high S/N ratio multicolored SNP detection technology toward each locus.

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A two-color readout of the Smart Amplification (SmartAmp、SMAP) genotyping system using so-called Exciton Primers have been developed. New method is highly specific and sensitive, permitting a one-step, single-tube mutation detection reaction which delivers a result in 45 minutes directly from blood. Exciton Primers, which are functioning as "fluorescence emission with sequence-specific," after hybridization to complementary sequences, can provide the signals resulting target sequence detection for real-time monitoring of amplification reactions. Applied to the isothermal SmartAmp2 mutation detection process, Exciton Primers show high signal strength with low background leading to a superior specificity and sensitivity compared to SYBR Green I. The genotyping assay can use only one labeled Exciton Primer for endpoint detection, or simultaneously by real-time monitoring detect wild-type and mutant alleles in a one-tube reaction using two Exciton Primers having different dyes. The technique seems very promising for decentralized point-of-care diagnostics since the approach does not involve complicated instrumentation and sample purification. We demonstrated efficiency of the new technology on the example of rapid detection of SNPs and mutations