HotStarTaq Plus Master Mix Kit

快速、高特异性扩增，适合多种应用

该产品将于 2023 年 11 月 30 日停产。

立即切换到后续 AllTaq PCR 试剂盒。了解有关过渡的更多信息，并查看详细的常见问答。

HotStarTaq Plus Master Mix Kit (1000)

Cat. No. / ID: 203645

For 1000 x 20 μl reactions: 12 x 0.85 ml HotStarTaq Plus Master Mix (contains 1000 units of HotStarTaq Plus DNA Polymerase, PCR Buffer with 3 mM MgCl₂, and 400 μM of each dNTP), 4 x 0.55 ml CoralLoad Concentrate, 8 x 1.9 ml RNase-Free Water

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HotStarTaq Plus Master Mix Kit 旨在用于分子生物学应用。该产品不能用于疾病诊断、预防和治疗。

该产品将于 2023 年 11 月 30 日停产。

立即切换到后续 AllTaq PCR 试剂盒。了解有关过渡的更多信息，并查看详细的常见问答。

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特点

快速激活热启动酶，只需5分钟
更少的移液操作，降低被污染风险
高PCR特异性，所需优化次数少
可直接上样的缓冲液，操作更简单

产品详情

HotStarTaq Plus Master Mix包含HotStarTaq DNA Polymerase、可将优化需求最小化的独特的QIAGEN PCR Buffer、以及dNTPs。HotStarTaq Plus Master Mix Kit与HotStarTaq Master Mix Kit一样，具备高特异性和高灵敏度，同时提供5分钟快速启动的聚合酶。预混液中提供所有组分，减少移液步骤、降低污染风险，同时提高通量和可重复性。此外，提供包含两种凝胶示踪染料的CoralLoad Concentrate，提高移液的可见性，并可直接将PCR产物上样至凝胶。

绩效

每一批HotStarTaq Plus Master Mix Kit都经过全方位的质量控制测试，包括：严格的PCR特异性和具有可重复性的试剂，即使低拷贝的靶分子也可扩增。通过检测，HotStarTaq DNA Polymerase可确保高度特异性和在热启动PCR中的卓越表现（参见" Highest specificity"和" Higher specificity with different primer–template systems"和表格）。该试剂盒提供的新型PCR缓冲液使得在多种PCR条件下都维持高特异性，无需优化。该试剂盒提供的CoralLoad Concentrate，提高移液的可见性，并可直接将PCR产物上样至凝胶，提高便利性。CoralLoad Concentrate可以添加到PCR反应物中，不会影响扩增的灵敏度和特异性。已经证实，在含有CoralLoad Concentrate的情况下可顺利扩增PCR片段，用于克隆和限制性酶切，无需预先纯化。

高度特异性配合简单的操作使HotStarTaq Plus Master Mix Kit适用于复杂的基因组或cDNA模板（参见" Effect of hot start on RT-PCR performance"）、多重引物对（参见" Specific amplification in multiplex PCR"）、从珍贵样本或低拷贝靶分子中抽提的模板（参见" Highly sensitive single-cell PCR"）。该试剂盒还适用于大量样本的扩增，诸如基因筛查等项目。

热启动模式比较
	HotStarTaq Plus DNA Polymerase	HotStarTaq DNA Polymerase	Supplier A_II提供的热启动酶	供应商R	供应商I （抗体介导）	手动	石蜡屏障
特异性扩增	++	++	+	++	+	+/–	+/–
PCR优化需求	++	++	+/–	+/–	+/–	–	–
易用性	+++	++	++	+	+	–	–
激活速度	++	+	++	++	++	–	–

HotStarTaq Plus DNA Polymerase特性

浓度：5 units/µl
重组酶：是
底物类似物：dNTP、ddNTP、dUTP、biotin-11-dUTP、DIG-11-dUTP、fluorescent-dNTP/ddNTP
延伸速度：72°C条件下2–4 kb/min
酶半衰期：97°C条件下10 min，94°C条件下60 min
扩增效率：≥10⁵倍
5'–>3'外切酶活性：是
额外添加A：是
3'–>5' 外切酶活性：否
核酸酶污染：否
蛋白酶污染：否
RNases污染：否
自启活性：否

查看图表

Highest specificity.

Higher specificity with different primer–template systems.

Effect of hot start on RT-PCR performance.

Specific amplification in multiplex PCR.

Highly sensitive single-cell PCR.

原理

HotStarTaq Plus Master Mix是一种即用型预混液，包含HotStarTaq DNA Polymerase、QIAGEN PCR Buffer、MgCl₂和dNTPs。HotStarTaq Plus DNA Polymerase，具备同HotStarTaq DNA Polymerase一样的卓越表现，激活时间只需5分钟。

HotStarTaq Plus DNA Polymerase是一种经修饰的QIAGEN Taq DNA Polymerase，以非活性形式提供，常温下没有聚合酶活性。避免PCR构建和循环初始阶段低温条件下非特异性引物的延伸和引物二聚体的形成（参见" Highest specificity"和" Higher specificity with different primer-template systems"）。95°C条件下孵育5分钟即可激活HotStarTaq Plus DNA Polymerase，这个步骤可整合入已有的热循环程序中。

QIAGEN PCR Buffer

QIAGEN PCR Buffer通过提高每个PCR扩增退火过程特异性引物的结合比例，确保PCR每个循环特异性扩增（参见" Increased specificity of primer annealing"）。独特的KCl和(NH₄)₂SO₄平衡组合，使得该缓冲液与传统的PCR缓冲液相比在很宽的温度和Mg²⁺浓度范围内都有严格的引物退火条件和高度的特异性，无需进行耗时的优化。

CoralLoad PCR Buffer

HotStarTaq Plus DNA Polymerase同时提供CoralLoad PCR Buffer，CoralLoad PCR Buffer具备QIAGEN PCR Buffer的所有优点，同时可直接将PCR产物上样到琼脂凝胶上，无需加入凝胶上样缓冲液。CoralLoad PCR Buffer具备同样高的PCR特异性，与常规的QIAGEN PCR Buffer一样可减少优化过程。此外，它包含两种指示染料，橙色染料和红色染料，可方便的判断DNA的迁移距离和优化跑胶时间（参见" CoralLoad PCR Buffer"）。该缓冲液提高移液的可见性，并可直接将PCR产物上样至凝胶，提高便利性。

查看图表

Highest specificity.

Higher specificity with different primer–template systems.

Increased specificity of primer annealing.

CoralLoad Concentrate.

程序

HotStarTaq Plus Master Mix Kit以方便的预混液形式提供，易于使用。95°C条件下孵育5分钟即可激活HotStarTaq Plus DNA Polymerase，可以整合入已有的热循环程序。在室温下应用该预混液可快速、简单地构建反应体系，只需将10 µl HotStarTaq Plus Master Mix加入每个PCR反应管，并加入10 µl溶于该试剂盒提供的RNase-free蒸馏水的引物和模板DNA（参见""）。移液步骤最少化，降低操作误差和污染风险，确保提高通量和可重复性。该试剂盒提供高效、经优化的实验方案，用于快速、方便的PCR反应体系构建。同时试剂盒提供CoralLoad Concentrate，提高移液的可见性，并可直接将PCR产物上样至凝胶，提高便利性。

查看图表

HotStarTaq Plus procedure.

应用

HotStarTaq Plus Master Mix Kit适合多种应用，包括各种富有挑战性的研究，如各种扩增应用：

复杂的基因模板
复杂的cDNA模板（如RT-PCR）
低拷贝的靶分子（如单细胞PCR）
多重引物对反应

辅助数据和图表

CoralLoad Concentrate.

Specifications

Features	Specifications
Applications	PCR, RT-PCR, Complex genomic templates, very low-copy targets
Enzyme activity	5' -> 3' exonuclease activity
Sample/target type	Genomic DNA and cDNA
Single or multiplex	Single
Real-time or endpoint	Endpoint
Mastermix	Yes
With/without hotstart	With hotstart
Reaction type	PCR amplification

资源

Second edition — innovative tools

Addressing critical factors and new solutions

For highly specific hot-start PCR without optimization

Download Safety Data Sheets for QIAGEN product components.

FAQ

HotStart PCR is a technique commonly used to improve the sensitivity and specificity of PCR amplifications. Lack of sensitivity or specificity is most often caused by the amplification of nonspecific priming events, such as primer dimers, that usually occur at the lower temperatures when reactions are set up. Although thermostable DNA-dependent DNA polymerases have optimal activity at higher temperatures, they do also have some activity at lower temperatures when they may amplify these nonspecific priming events. HotStart enzymes are inactive at room temperature, and require heating at nucleic acid melting temperatures in order to be activated. In this way, nonspecific priming events are melted before the enzyme can amplify them. During PCR cycles, the temperature never drops low enough during annealing of gene-specific primers for nonspecific priming events to occur, resulting in amplification exclusively of the target of interest. When using a HotStart DNA polymerase, it is critical that the initial denaturation step in the experiment be of sufficient duration to fully activate the enzyme.

FAQ ID -2676

Yes, please follow the Supplementary Protocol 'Polyacrylamide_gel_analysis_of_oligonucleotides' (PCR03).

FAQ ID -961

Not necessarily. In a lot of cases, the uniquely formulated PCR Buffer provided in the HotStarTag Plus DNA Polymerase, HotStar HiFidelity Polymerase, Taq DNA Polymerase, HotStarTaq DNA Polymerase, and QIAGEN Multiplex PCR Kits provides optimal amplification of specific PCR products. The usefulness of Q-Solution needs to be determined empirically for each primer/template setup, by running parallel PCR reactions with and without Q-Solution under the same cycling conditions.

Q-Solution changes the melting behavior of DNA and will often improve a suboptimal PCR caused by templates that have a high degree of secondary structure or high GC-contents. For more details on the effects of Q-Solution on PCR amplification, please see the Q-Solution sections of the HotStarTaq Plus DNA Polymerase, HotStar HiFidelity Polymerase, Taq DNA Polymerase, HotStarTaq DNA Polymerase, and the QIAGEN Multiplex PCR Handbooks.

FAQ ID -380

CoralLoad gel tracking dye contained in Taq, HotStarTaq, TopTaq DNA Polymerase and TopTaq Master Mix Kits separates into 2 fragment-size dependent colors (orange and red) when loaded onto an agarose gel. Sigma Red buffer only has one color which is harder to visualize.

FAQ ID -1644