TopTaq Master Mix Kit

高度可靠的终点法PCR，无可比拟的方便性

Features

易于使用的预混液，操作步骤最少化
冷藏储存节省了解冻时间
可靠的高产率PCR
方便的室温操作
优化的操作程序适合所有PCR实验

Product Details

TopTaq Master Mix Kit结合了TopTaq DNA Polymerase和即用型预混液的所有优势，简单易用，包含TopTaq DNA Polymerase，将优化需求最小化的TopTaq PCR Buffer和dNTPs。酶保存缓冲液中有独特的TopTaq稳定剂，使得TopTaq Master Mix可保存在4°C，并在室温下进行反应，无需耗时的解冻试剂。所有的成分都含在即用型混合液中，减少了移液步骤，降低了污染的风险。TopTaq Master Mix即使在25°C或4°C下存放很长一段时间，还是可以获得高产率的PCR产物。

Performance

TopTaq Master Mix Kit用于标准的和特有的终点式PCR应用，无需优化。TopTaq Master Mix即使在25°C或4°C下保存4个月还是可以获得高产率的PCR产物，说明TopTaq Master Mix能够提供非常稳定和可靠的扩增体系（参见""）。TopTaq稳定剂与TopTaq DNA Polymerase结合，防止聚合酶在长期储存过程中灭活。预混液中含有独特的PCR，具有均衡的钾和铵离子，改进特异性引物-模板的退火条件，并同时降低非特异性退火条件。TopTaq DNA Polymerase比其他供应商测试的DNA聚合酶具有更好的性能，确保了即使在使用较低的模板量，也可获得高产量的特异性产物和单独的片段大小（参见"”）。

See figures

Reliable high-yield PCR, independent of storage conditions.

High yields of PCR product without optimization

Principle

由TopTaq Master Mix Kit研发用于提供前所未有的应用便捷性和高度可靠的终点式PCR。在研发TopTaq PCR体系之前，所有PCR酶都需要在–20°C下储存。因为在酶保存缓冲液中含有独特的专有TopTaq稳定剂，TopTaq Master Mix Kit是首款可在4°C下储存的试剂盒。该即用型预混液规格提供了更大的便利。由于TopTaq DNA Polymerase的使用，甚至在扩增不同大小的片段时，也可获得高产量的特异性产物。（参见" High PCR product yields without optimization"）。

TopTaq PCR Buffer

新型的TopTaq PCR Buffer有助于扩增特异性PCR产物。在每个PCR循环的退火步骤中，相对非特异性引物结合，该缓冲液大大增加了特异性引物结合的比率。由于KCl和(NH₄)₂SO₄独特均衡的结合，相比传统的PCR缓冲液，该PCR缓冲液提供了更广泛的温度和Mg²⁺浓度，以及严格的引物-退火条件（参见" Increased specificity of primer annealing"）。通过调节退火温度或Mg²⁺浓度可显著减少优化PCR反应的需求，有时不需要优化过程。

TopTaq稳定剂

TopTaq与TopTaq DNA Polymerase结合，防止在长期储存过程中聚合酶灭活（参见" Stabilization at 4°C"），同时防止模板DNA和引物在低温下与聚合酶结合。在初始的降解步骤中，TopTaq稳定剂能从聚合酶中分离而不影响聚合酶的活性。

CoralLoad Concentrate

为了更方便使用，成套提供TopTaq Master Mix Kit和CoralLoad Concentrate。CoralLoad Concentrate包含两种胶示踪染料。该独特的试剂提高了移液可视化，有助于去除DNA迁移距离并优化琼脂糖凝胶跑样时间。使用CoralLoad Concentrate时，可将PCR产物直接上样到琼脂糖凝胶上而不需要事先另外加缓冲液（参见" Convenient CoralLoad Concentrate"）。

See figures

High yields of PCR product without optimization

Increased specificity of primer annealing.

Stabilization at 4°C.

Convenient CoralLoad Concentrate.

Procedure

TopTaq Master Mix Kit结合了独特的TopTaq DNA Polymerase和即用型预混液的所有优势。由于TopTaq Master Mix Kit，无需单独的移液部分组件，并能最大限度地减少污染风险和移液可视化，使PCR反应体系构建快速简单。在初始的降解步骤中，TopTaq稳定剂能从聚合酶中分离而不影响聚合酶活性。这增加了便利，并能即可进行反映设置，无需冰上工作，节省了解冻试剂所需的时间（参见""）。

可靠的高产量PCR，无需优化过程

经优化的实验方案适用于所有PCR检测，去除了个别引物-模板体系中漫长的实验参数优化需要，能节省大量的时间和成本。TopTaq Master Mix即使在25°C或4°C下保存4个月还是可以获得高产率特异性PCR产物（参见" Reliable high-yield PCR"）。

See figures

TopTaq procedure.

Reliable high-yield PCR, independent of storage conditions.

Applications

TopTaq Master Mix Kit高度适用于所有终点法PCR应用。

Supporting data and figures

Reliable high-yield PCR, independent of storage conditions.

PCR was performed using TopTaq Master Mix stored at –20°C, 4°C, and 25°C for 4 months. Two different genes from human genomic DNA (prp, 750 bp and HugL, 1200 bp) were amplified according to the standard optimized protocol (duplicates shown). The results show high yields of specific PCR product. [A] Gel image produced by the QIAxcel System. Markers: GelPilot 1 kb Plus Marker and QX Alignment Marker 15 bp/10 kb. [B] The QIAxcel System fully automates sensitive, high-resolution capillary electrophoresis of up to 96 samples per run.

Specifications

Features	Specifications
Applications	PCR, RT-PCR, DNA Fingerprinting
Real-time or endpoint	Endpoint
dNTP's included	Yes (in Master Mix)
Mastermix	Yes
Enzyme activity	5' -> 3' exonuclease activity
Reaction type	PCR amplification
Sample/target type	Genomic DNA and cDNA
Single or multiplex	Single
Storage conditions	Fridge storage and room temperature setup
With/without hotstart	Without Hotstart

Resources

FAQ

Nucleotide analogs that can be used with TopTaq DNA Polymerase are:

dNTP
ddNTP
biotin-dATP
dITP

FAQ ID -1741

No. TopTaq DNA Polymerase is not a hot-start polymerase. Although TopTaq Stabilizer reduces enzyme accessibility at lower temperatures, it does not allow stringent hot-start procedures.

FAQ ID -1748

No. TopTaq Stabilizer contained in the enzyme storage buffer of the TopTaq DNA Polymerase and TopTaq Master Mix Kits does not interfere with downstream applications such as cloning, sequencing, or restriction digestion.

FAQ ID -1744

CoralLoad dyes supplied in PCR Kits such as, e.g., Taq, HotStarTaq, and TopTaq DNA Polymerase and TopTaq Master Mix do not interfere with most downstream enzymatic applications.

However, for reproducible results, purification of PCR products using the QIAquick or MinElute PCR Purification Kits prior to enzymatic manipulation is recommended.

FAQ ID -1745

Our data show stability of the TopTaq Master Mix Kit at a constant temperature of 25°C for up to 4 months. However, we do not recommend storage of TopTaq Master Mix on the bench due to temperature fluctuations typical in standard laboratories; nor can we guarantee the performance of TopTaq Master Mix under this storage condition.

FAQ ID -2052

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

TopTaq DNA Polymerase and the TopTaq Master Mix Kit are shipped on dry ice but retain full activity at room temperature (15-25°C) for at least 2 weeks. Both kits can be stored at 2-8°C for up to 12 months.

FAQ ID -1743

The maximum amplicon length we have tested using TopTaq DNA Polymerase and the TopTaq Master Mix Kit was 5 kb, amplified from genomic DNA template. Larger PCR amplicon sizes are possible when using less complex template DNA.

FAQ ID -1738

The error rate of TopTaq DNA Polymerase is very similar to that of standard Taq DNA Polymerase: approximately 2-3x 10e-5 (per base, per cycle).

FAQ ID -1739

Yes. The terminal transferase activity of TopTaq DNA Polymerase adds a 3`A-overhang to the PCR product and therefore enables TA- and UA- cloning.

FAQ ID -1740