Hot Start rTth (DNA) Kit is qPCR reagent based on Tth DNA Polymerase. Tth DNA Polymerase has higher amplification efficiency than Taq DNA Polymerase, which is a general-purpose enzyme, and enables amplification from low copies of template and amplification from a crude sample containing PCR inhibitors.
In addition, Tth DNA Polymerase has a 5 '- 3' exonuclease activity, so it can be used for real-time PCR using probe assays such as TaqMan® assay. This enzyme contains neutralizing antibodies, thus allowing for Hot start PCR.
- Excellent DNA Amplification Efficiency
The reaction composition is optimized based on Tth DNA Polymerase, enabling efficient PCR even from low copies of template.
- Tolerant of PCR Inhibitors
This kit is effective for amplification from crude samples (e.g., biological samples, foodstuffs, soil extract, etc.). In the case of amplification from whole blood, sufficient amplification can be achieved by adding it directly to the reaction solution without purification of nucleic acid.
- Utilization of dUTP
This kit contains dUTP instead of dTTP in 2x Buffer for rTth/ TTx (DNA). Therefore, the rate of false-positive detection can be reduced by adding Uracil-N-glycosylase (UNG).
*UNG is not supplied with this kit. Uracil-DNA Glycosylase (UNG), Heat-labile(Code No. UNG-101) can be used.
TaqMan® assays or hybridization probe assays using DNA template
Store at -20℃
This kit includes the following components for 250 reactions, 20 μL total reaction volume. All reagents should be stored at -20°C.
|2x Buffer for rTth/ TTx (DNA)||1.25 mL x 2|
|Hot Start rTth DNA Polymerase (4U/ μL)||62.5 μL|
2× Reaction Buffer contains essential components for the reaction (buffer, salts, Mg2+, dATP, dCTP, dGTP, and dUTP, etc.). Add template DNA, primers, and attached Hot Start rTth DNA Polymerase, and adjust to 1x concentration with sterile water etc.
DNA Polymerase is a mixture of rTth DNA polymerase and hot start antibodies. Its concentration is 4U/ μL.
This kit doesn’t contain a passive reference dye (ROX). When using a passive reference dye to compensate fluorescence intensity and dispensing error between wells, please use the separately sold 50x ROX reference dye (Code No. ROX-101).
Example 1.Detection of SNPs
SNPs in aldehyde dehydrogenase 2 (ALDH2) gene were detected using TaqMan® probes.
1 μL of whole blood was added to 20 μL reaction mixture as template. As a result, Taq DNA Polymerase-based reagent was inhibited by whole blood and amplification could not be confirmed. On the other hand, Hot Start rTth (DNA) Kit was able to determine SNPs from whole blood without purification.