Photoinduced Electron Transfer Fluorescent Primer for Nucleic Acid Amplification

CDC scientists have developed a rapid and cost-efficient method for generating fluorescently labeled primers for PCR and real-time PCR. At present, fluorescent primers are useful for detecting and identifying microbes and specific nucleic acid sequences, amplifying nucleic acids for pyro-sequencing, determining the levels of gene expression, and many other uses. However, problems exist with current techniques used to create fluorescent primers. For one, labeling is not one hundred percent efficient, leading to inaccurate results. Further, it is expensive and time consuming for researchers to make and label their own unique primers. This technology allows for the creation of custom primers in which fluorescent dye attaches to all oligomers.

This technology employs photoinduced electron transfer (PET) nucleic acid molecules that can be used detect and amplify target nucleic acid molecules. PET tags are attached to the 5'-end of a target-specific oligo for fluorescent labeling of the primer. PET tag activity can be quenched by at least two consecutive guanosines (G-G) within the tag sequence and activity is un-quenched when the PET tag hybridizes with its complementary nucleic acid molecule.