For general laboratory use. Others LightCycler RNA Amplification Kit HybProbe GLU LightCycler® RNA Amplification Kit HybProbe 3.5.5.1.2.2 12015145001 LC RNA Amplification Kit, Hyb. Probes LightCycler RNA Amplification Kit HybProbe 04038377016450 Reagents, kits 1 kit 96 reactions of 20 μl final volume each false Easy-to-use Reaction Mix for One-Step RT-PCR, using HybProbe probes with the LightCycler® Carousel-Based System. en HybProbe probes are two different short oligonucleotides that bind to an internal sequence of the amplified fragment during the annealing phase of the amplification cycle. One probe is labeled at the 5' end with a fluorophore dye (LightCycler® Red 610, 640, Cy5, or Cy5.5); it is also 3'-phosphorylated so it cannot be extended. The other probe is labeled at the 3' end with fluorescein. When hybridized to the template DNA, the two probes are close enough to allow fluorescence resonance energy transfer (FRET) between the two fluorophores.During FRET, fluorescein (the donor fluorophore) is excited by the light source of the LightCycler® Instrument. Fluorescein transfers part of this excitation energy to the dye (the acceptor fluorophore). Then, the dye emits fluorescence which is measured by the LightCycler® Instrument. HybProbe probes  that contain different dye labels can be used separately for single-color detection experiments, or combined for dual-color detection experiments.Color compensation is not necessary for single-color detection experiments. However, if you are using HybProbe probes to perform dual-color experiments in a single capillary, you must also use a color compensation file. Color compensation may be applied either during or after a run on the LightCycler® Instrument.LightCycler® Red 610 and Cy5 can be used only on a LightCycler® 2.0 Instrument.How this Product WorksThe LightCycler® RNA Amplification Kit HybProbe is designed specifically for the HybProbe detection format using the LightCycler® Carousel-Based System. It is used to perform one-step RT-PCR in 20 μl glass capillaries. Amplification and online monitoring of the template RNA is achieved by a combined procedure on the LightCycler® Instruments. The results are interpreted directly after completing the PCR. The amplicon is detected by fluorescence using target-specific HybProbe probes (not supplied with the kit).The LightCycler® RNA Amplification Kit HybProbe provides convenience, high performance, reproducibility, and minimizes contamination risk. Only template RNA, PCR primers, HybProbe probes, and additional MgCl2 (if necessary), need to be added.In principle, the LightCycler® RNA Amplification Kit HybProbe can be used for the amplification and detection of every RNA target. However, you would need to adapt each amplification protocol to the reaction conditions of the LightCycler® Carousel-Based System and design specific PCR Primers and HybProbe probes for each target. See the LightCycler® Operator's Manual for general recommendations.​​​​​​​The amplicon size should not exceed 1 kb in length. For optimum results, select a product length of 700 bp or less. en The LightCycler® RNA Amplification Kit HybProbe is designed specifically for the HybProbe detection format using the LightCycler® Carousel-Based System. The kit provides reagents, including RT-PCR enzyme mix, reaction mix, MgCl2, and Water, PCR Grade for very sensitive detection and quantification of defined RNA sequences using the LightCycler® System (suitable RT-PCR primers and HybProbe probes must be supplied). It can also be used to genotype single nucleotide polymorphisms (SNPs) and analyze mutations.In addition, the kit can be used with Uracil-DNA Glycosylase (UNG), heat labile to prevent carryover contamination during PCR. en