Description
The polymerase combines the high specificity, sensitivity, and minimal optimization of HotStarTaq DNA Polymerase, with a fast 5-minute activation time. PCR can be set up at room temperature and reactions can be directly loaded onto a gel, due to novel CoralLoad PCR Buffer, which contains two gel-tracking dyes. The standard QIAGEN PCR Buffer is also included for greater convenience. In addition, Q-Solution, a novel additive that enables efficient amplification of "difficult"(e.g., GC rich) templates, is also provided. The unique kit components and optimized protocols streamline the PCR procedure.
Each lot of HotStarTaq Plus DNA Polymerase is subjected to a comprehensive range of quality control tests, including a stringent PCR specificity and reproducibility assay in which low-copy targets are amplified. HotStarTaq Plus DNA Polymerase outperformed kits tested from other suppliers and ensures high specificity and superior performance in hot-start PCR (see figures "Highest specificity "and "Higher specificity with different primer–template systems", and table). The innovative PCR buffer provided with the kit ensures specificity over a wide range of PCR conditions, minimizing the need for optimization. Suboptimal PCR can be improved with Q-Solution, also provided with the kit (see figure "Amplification of difficult templates"). Together, these components ensure specific amplification in a range of applications (see figure "Effect of hot start on RT-PCR performance "and "Highly sensitive single-cell PCR"). Comparison of hot-start methods HotStarTaq Plus DNA Polymerase HotStarTaq DNA Polymerase Hot-start enzyme from Supplier A II Supplier R Supplier I (antibody-mediated) Manual Wax barrier Specific amplification ++ ++ + ++ + +/– +/– Minimal PCR optimization ++ ++ +/– +/– +/– – – Easy to use +++ ++ ++ + + – – Speed of activation ++ + ++ ++ ++ – – HotStarTaq Plus DNA Polymerase specifications Concentration: 5 units/µl Recombinant enzyme: Yes Substrate analogs: dNTP, ddNTP, dUTP, biotin-11-dUTP, DIG-11-dUTP, fluorescent-dNTP/ddNTP Extension rate: 2–4 kb/min at 72°C Half-life: 10 min at 97°C; 60 min at 94°C Amplification efficiency: ≥10 5 fold 5'–>3' exonuclease activity: Yes Extra A addition: Yes 3'–>5' exonuclease activity: No Contaminating nucleases: No Contaminating RNases: No Contaminating proteases: No Self-priming activity: No
HotStarTaq Plus DNA Polymerase provides the unrivaled performance of HotStarTaq DNA Polymerase with a shortened activation time of just 5 minutes. HotStarTaq Plus DNA Polymerase, a modified form of QIAGEN Taq DNA Polymerase, is supplied in an inactive state that has no polymerase activity at ambient temperatures. This prevents extension of nonspecifically annealed primers and primer dimers formed at low temperatures during PCR setup and the initial PCR cycle (see figures "Highest specificity "and "Higher specificity with different primer-template systems"). HotStarTaq Plus DNA Polymerase is activated by a short 5-minute incubation at 95°C which can be easily incorporated into any existing thermal-cycler program. QIAGEN PCR Buffer QIAGEN PCR Buffer maintains specific amplification in every cycle of PCR by promoting a high ratio of specific-to-nonspecific primer binding during the annealing step in each PCR cycle (see figure "Increased specificity of primer annealing"). Owing to a uniquely balanced combination of KCl and (NH 4) 2 SO 4, the buffer provides stringent primer-annealing conditions over a wider range of annealing temperatures and Mg 2+ concentrations than conventional PCR buffers. Optimization of PCR by varying the annealing temperature or the Mg 2+ concentration is therefore often minimal or not required. CoralLoad PCR Buffer HotStarTaq Plus DNA Polymerase is supplied with CoralLoad PCR Buffer, which has all of the advantages of QIAGEN PCR Buffer but can also be used to directly load the PCR reaction onto an agarose gel without the need to add a gel loading buffer. CoralLoad PCR Buffer provides the same high PCR specificity and minimal reaction optimization as the conventional QIAGEN PCR Buffer. Additionally, it contains two marker dyes — an orange dye and a red dye — that facilitate estimation of DNA migration distance and optimization of agarose gel run time (see figure "CoralLoad PCR Buffer"). The buffer ensures improved pipetting visibility and enables direct loading of PCR products onto a gel, for enhanced convenience. Q-Solution Q-Solution, an innovative PCR additive that facilitates amplification of difficult templates by modifying the melting behavior of DNA, is also provided with HotStarTaq DNA Polymerase. This unique reagent improves suboptimal PCR caused by templates that have a high degree of secondary structure or GC-rich templates (see figure "Amplification of difficult templates"). Unlike other commonly used PCR additives such as DMSO, Q-Solution is used at just one working concentration, is nontoxic, and PCR purity is guaranteed. Adding Q-Solution to the PCR does not compromise PCR fidelity.
HotStarTaq Plus DNA Polymerase is supplied with a streamlined, optimized protocol for fast and easy PCR setup. HotStarTaq Plus DNA Polymerase is activated by a 5-minute, 95°C incubation step, which can easily be incorporated into existing thermal cycling programs. Reactions can be set up at room temperature, ensuring greater convenience and ease of use (see figure "HotStarTaq Plus procedure"). CoralLoad PCR Buffer, also provided with the kit, ensures improved pipetting visibility and enables direct loading of PCR products onto a gel for enhanced convenience.
