Design and Development of a Telomere Sensor Based on Fluorescence Energy Transfer

Abstract

A 200 nm in-diameter single-bead sensor for detecting single unlabeled DNA molecules in solution based on fluorescence resonance energy transfer is presented. Both bulk and single molecule fluorescence experiments were carried out at room temperature. The G-rich telomere forming a G-quadruplex structure in the presence of ZnTCPP showed two strong donor and acceptor signals. DNA-bound Alexa 488 and crimson 625 loaded in a commercial bead serve as the donor and acceptor, respectively. When Binding of the target DNA to the single bead sensor induces G-quadruplex stretching, resulting in a decrease in fluorescence energy transfer. The limit of sensor's sensitivity can reach 1 fM. Under optimized conditions using a Polydimethylsiloxane (PDMS) microfluidic device direct bead-sensor counting is used to measure the number of beads by control of the flow rate through the probe volume and finish one experiment in 5 minutes. A new parameter based on the ratio of donor and acceptor fluorescence signal from 0.1 to 100 was proposed and used to define the sensitivity and selectivity of bio-sensor.