Hot Start Taq DNA Polymerase is a hot-start Taq enzyme obtained by mixing Taq antibody and Taq DNA polymerase in a certain proportion, which has higher stability and detection rate. Based on the thermal stability of Taq antibody, Hot Start Taq DNA Polymerase can still maintain strict sealing at 45℃, so that non-specific amplification is inhibited during the mixed sample and system heating stage. When the reaction is maintained at 95℃ for more than 30 seconds, Taq antibody is inactivated, and the activity of Taq enzyme is completely released, ensuring that the PCR system has high amplification sensitivity and specificity. Hot Start Taq DNA Polymerase activation is not affected by factors such as buffer pH and ionic strength, and is suitable for various hot-start PCR and qPCR reactions based on Taq DNA polymerase. It is commonly used to amplify low-copy genes from complex templates (genome, cDNA), and is based on PCR/qPCR molecular diagnostic reagents using hot-start Taq enzyme.

UA070096: 5 U/μL Hot Start Taq DNA Polymerase, Taq antibody, 20 mM Tris-HCl (pH 8.0@ 25°C), 100 mM KCl, 1 mM DTT, 0.1 mM EDTA, 0.5% (v/v) Nonidet P40, 0.5% (v/v) Tween 20 and 50% (v/v) glycerol
10* Reaction Buffer: 100mM Tris-HCl (pH9.0 @25℃), 500mM KCl, 1% Triton® X-100,15mM MgCl2。

a. Dissolve and mix the various solutions required for the PCR reaction. Due to the hot start nature of the enzyme, reactions can be assembled on the bench at room temperature and transferred to a thermocycler. No separate activation step is required to release the antibody from the enzyme.
b. Refer to the table below to set up the PCR reaction system on the ice bath (if there are multiple similar PCR reactions, you can first prepare a large volume mixture containing water, buffer, dNTP, and Hot Start Taq DNA Polymerase, and then divide it into each PCR reaction tube. According to the situation, sometimes the mixture can include primers):

Notes: Gently mix the reaction. Collect all liquid to the bottom of the tube by a quick spin if necessary. Overlay the sample with mineral oil if using a PCR machine without a heated lid.
c. Transfer PCR tubes from ice to a PCR machine with the block preheated to 95°C and begin thermocycling.
Thermocycling conditions for a routine PCR:

1. Since the PCR reaction is very sensitive and can amplify the target gene sequence more than 10 million times, please pay attention to avoid contamination of the small amount of DNA to be amplified when using Hot Start Taq Polymerase, and try to consider setting a blank control without a template to confirm whether there is contamination of the DNA to be amplified.

2. 2mM Mg2+ can meet the needs of most PCR amplification, and for some PCR, it can be adjusted to 2-4mM to ensure better amplification.