LightCycler probes are highly sensitive and sequence-specific fluorescent probes designed for use with the Roche LightCycler instruments.
Several reporters are available and are suitable for multiplex analysis.
A LightCycler Probe system consists of a pair of single-stranded fluorescent-labeled oligonucleotides. Oligo Probe 1 is labeled at the 3’ end with a donor reporter and Oligo Probe 2 is labeled at its 5’ end with one of a few available acceptor reporters. The free 3’ hydroxyl group of Probe 2 must be blocked with a phosphate group (P) to prevent DNA polymerase extension. There should be a space of 1 to 5 nt to separate the two probes from each other. The structures and mechanism of a LightCycler probe are shown below.
During the annealing step of qPCR, the PCR primers and the LightCycler Probes hybridize to their specific target regions bringing the donor dye into close proximity to the acceptor dye. When the donor dye is excited by light from the LightCycler instrument, energy is transferred by FRET from the donor to the acceptor dye. The acceptor reporter’s emission wavelength is detected. The increase in fluorescence signal is directly proportional to the amount of target DNA.
Our probes are provided in a format to simplify your experimental planning.
The recommended constructs for Oligo Probe 1 and Oligo Probe 2 are listed in the tables below: