Method Description

This polymerase chain reaction (PCR) method employs a target-specific detection system, including primers and fluorescent resonance energy transfer (FRET) hybridization probes designed for the ureC gene of Ureaplasma urealyticum and Ureaplasma parvum. The LightCycler instrument amplifies and monitors target nucleic acid sequences by fluorescence during PCR cycling. This is an automated PCR system that can rapidly detect amplified product development. The detection of amplified products is based on the FRET principle. For FRET product detection, a hybridization probe with a donor fluorophore, fluorescein, on the 3' end is excited by an external light source, which emits light that is absorbed by a second hybridization probe with an acceptor fluorophore, LC-Red 640, on the 5' end. The acceptor fluorophore then emits light of a different wavelength that is measured with a signal that is proportional to the amount of specific PCR product. The process is completed in a closed tube system and the melting temperature of the probes allows differentiation of U urealyticum from U parvum.(Cunningham SA, Mandrekar JN, Rosenblatt JE, Patel R: Rapid PCR Detection of Mycoplasma hominis, Ureaplasma urealyticum, and Ureaplasma parvum. Int J Bacteriol 2013 Jan 30, doi: 10.1155/2013/168742)