Multiplex Single-Nucleotide Polymorphism Detection by Combinatorial Fluorescence Energy Transfer Tags and Molecular Affinity

Combinatorial fluorescence energy transfer (CFET) tags, constructed by exploiting fluorescence energy transfer and combinatorial synthesis to generate a large number of unique fluorescence emission signatures from a limited number of fluorophores, allow multiple biological targets to be identified simultaneously. All of the CFET tags can be excited by a single wavelength of 488 nm and analyzed by a simple optical system. In genetic analysis, the CFET tags are coupled with solid phase capture for multiplex singlenucleotide polymorphism (SNP) detection. The design, synthesis, purification of CFET tags, and the methods to use the CFET tags and molecular affinity for SNP detection in the retinoblastoma tumor suppressor gene are described.