EAGER: BISH: Isothermal solid-phase PCR using electrostatic control of DNA hybridization

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0939048VainrubThe proposed research investigates the hitherto unknown fundamental characteristics of electrostatic DNA hybridization. Namely; the yield, kinetic rates, ability to denature long DNA and to support a sufficient number of hybridization cycles, in order to establish conditions for efficient PCR amplification. The specific aims are to: 1. Construct a fluorescent detector for hybridization and extension of electrode tethered primers.An evanescent wave fluorescent real-time detector will be built to measure the kinetics and yieldof denaturation, annealing, and extension stages in solid-phase PCR.2. Prove-in-principle and optimize electrostatic isothermal PCR. Linear and exponentialamplification will be demonstrated in real-time PCR mode using oligonucleotide templates.Efficient electrostatic pulse cycles and optimal surface and solution conditions will be defined.Amplification protocols will be developed.