We now offer a complete diagnostics set for the 4 sexually transmitted urinal infections. Chlamydia trachomatis, Neisseria gonorrhoeae, Ureaplasma sp., Mycoplasma hominis / genitalium
EliGene kits and Real-Time PCR instruments
EliGene® kits are divided according to detection chemistry to two basic lines, RT and LC respectively.
In RT line, the TaqMan chemistry using hydrolyzing fluorescence probes is used in the combination with hot-start polymerase and ROX dye as a passive reference (see Figure 1A). The total reaction volume is 25ul when 5ul of isolated DNA is added directly to ready-to use mastermix containing all necessary components for reaction. In RT kits; the fluorescence dyes with excitation wavelengths of 495, 520 nm and emission wavelengths of 520, 550 nm together with non-fluorescence quenchers are used. These kits are basically designed for ABI instruments of 7000 line and are optimized on LightCycler480 instrument but can be used with other instruments that meet the spectral characteristics of used fluorescence dyes. The typical time of the run for RT kits is about 2 hours when list of compatible instruments you can find in Table 1.
In LC line, the Molecular Beacons chemistry using hybridization fluorescence probes is used in combination with hot-start polymerase (see Figure 1B). This line of kits is further divided into two versions, LC and LC2 that differs by used fluorescence dyes. The total reaction volume is 20ul when 5ul of isolated DNA is added to mastermix created by mixing of polymerase with reaction mix containing all necessary components for reaction. In LC version of the kits, the fluorescence dyes with excitation wavelengths of 495, 460 nm and emission wavelengths of 530, 705 nm together with non-fluorescence quenchers are used while in LC2 version, the fluorescence dyes with excitation wavelengths of 495, 520 nm and emission wavelengths of 530, 550 nm together with non-fluorescence quenchers are used. These kits are basically designed for LightCycler 1.2 and 2.0 instruments, kits are optimized on LightCycler480 instrument but can be used with other instruments that meet on the one hand the spectral characteristics of used fluorescence dyes in individual versions and on the other hand having rapid heating/cooling rate. The typical time of the run for LC kits is about 1 hours when the list of compatible instruments you can find in Table 1.
Figure A. Basic principle of TaqMan hydrolization probes (A) and Molecular Beacons hybridization probes (B) action.
The TaqMan probe hybridizes between the primers in annealing step and during elongation step probe is cleaved by exonuclease activity of Taq Polymerase. The Molecular Beacon probe forms beacon-like structure that opens to specific sequence during the hybridization in annealing step (B). “TaqMan” is registered trademark of Roche Molecular Systems Inc., “Molecular Beacon” is registered trademark of Public Health Research Institute Properties, Inc. (PHRI)
