高度可靠的PCR实验,无可比拟的方便性

  • 4°C的储存条件,可立即进行反应体系构建
  • 方便的室温操作
  • 经优化的操作流程适合所有PCR实验
  • 特异性扩增,DNA产量高

TopTaq DNA Polymerase具有一系列方便易用的特点,提供前所未有的应用便捷性和高DNA产量。酶保存液中含有独特的TopTaq稳定剂,可以在4°C下稳定保存TopTaq DNA Polymerase,并且在室温下进行反应体系构建,无需费时的解冻过程。CoralLoad Concentrate包含两种胶示踪染料,便于移液可视化,并可将PCR产物直接上样电泳。即使在相同的Mg2+浓度和退火温度下扩增不同长度的片段,依然可获得高产量PCR产物。

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TopTaq DNA Polymerase (50)
50 units TopTaq DNA Polymerase (proprietary recombinant polymerase isolated from a thermophilic eubacterium), 10x PCR Buffer (contains 15 mM MgCl2), 10x CoralLoad Concentrate, 5x Q-Solution, 25 mM MgCl2
200201
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TopTaq DNA Polymerase (250)
250 units TopTaq DNA Polymerase (proprietary recombinant polymerase isolated from a thermophilic eubacterium), 10x PCR Buffer (contains 15 mM MgCl2), 10x CoralLoad Concentrate, 5x Q-Solution, 25 mM MgCl2
200203
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TopTaq DNA Polymerase (1000)
4 x 250 units TopTaq DNA Polymerase (proprietary recombinant polymerase isolated from a thermophilic eubacterium), 10x PCR Buffer (contains 15 mM MgCl2), 10x CoralLoad Concentrate, 5x Q-Solution, 25 mM MgCl2
200205
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TopTaq DNA Polymerase (5000)
5000 units TopTaq DNA Polymerase (proprietary recombinant polymerase isolated from a thermophilic eubacterium; supplied in a single vial), 10x TopTaq PCR Buffer (contains 15 mM MgCl2), 10x CoralLoad Concentrate, 5x Q-Solution, 25 mM MgCl2
200207
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The TopTaq DNA Polymerase is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

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Convenient CoralLoad Concentrate.|TopTaq procedure.|High yields of PCR product without optimization.|Increased specificity of primer annealing.|Stabilization at 4°C.|
[A] CoralLoad Concentrate contains two gel-tracking dyes, enabling immediate gel loading of PCR products for [B] easy visualization of DNA migration.|Due to the unique proprietary TopTaq Stabilizer contained in the enzyme storage buffer, TopTaq DNA Polymerase is the first PCR kit that can be stored at 4°C. TopTaq Stabilizer binds to TopTaq DNA Polymerase, preventing polymerase inactivation. Template DNA and primers are also prevented from binding to the polymerase at low temperatures. During the initial denaturation step, the TopTaq Stabilizer freely dissociates from the polymerase without compromising polymerase activity. This added convenience allows immediate reaction setup without the use of ice — saving the time usually required for reagent thawing.|[A] TopTaq DNA Polymerase was used to amplify 14 different PCR products from mammalian genomic DNA ranging in size from 100 bp to 2 kb according to the standard preoptimized protocol and using identical cycling conditions. [B] The same four targets (shown in lanes 11–14) were amplified under identical cycling conditions using DNA polymerases from the indicated suppliers. TopTaq DNA Polymerase (QIAGEN) resulted in high yields of specific PCR products, regardless of fragment size.|Ammonium and potassium cations in QIAGEN PCR Buffers increase specificity of primer annealing. K+ binds to the phosphate groups (P) on the DNA backbone, stabilizing the annealing of the primers to the template. NH4+, which exists both as the ammonium ion and as ammonia under thermal-cycling conditions, can interact with the hydrogen bonds between the bases (B), destabilizing the weak hydrogen bonds at mismatched bases. The combined effect of the two cations maintains a high ratio of specific-to-nonspecific primer-template binding over a wide temperature range.|[A] TopTaq DNA Polymerase is stable at 4°C or room temperature due to the presence of low-molecular-weight molecules (TopTaq Stabilizer) on the surface of the polymerase. [B] At higher temperatures, the TopTaq Stabilizer completely dissociates from the polymerase without compromising polymerase activity.|
Performance
TopTaq DNA Polymerase用于常规的和特殊的终点式PCR应用。TopTaq DNA Polymerase是第一款可以在4°C下储存的PCR试剂盒。TopTaq Stabilizer(包含在酶储存缓冲液中)结合TopTaq DNA Polymerase,防止在长期储存过程中聚合酶灭活。在初始的降解步骤中,TopTaq稳定剂从聚合酶中分离而不影响其活性。TopTaq DNA Polymerase比其他供应商测试的DNA聚合酶有更好的性能,甚至在使用低模板量时,也能确保特异性产物的高产量和独立的片段大小(参见"High PCR product yields without optimization“)。
Principle

TopTaq DNA Polymerase是重组的94 kDa耐热DNA聚合酶。由QIAGEN研发的TopTaq DNA Polymerase,提供前所未有的应用便捷性和高度可靠的终点式PCR。直到研发了TopTaq PCR体系,所有PCR酶都需要在–20°C储存;然而,因为酶保存液中有独特的专有TopTaq稳定剂,TopTaq DNA Polymerase是第一款能在4°C下储存的试剂盒。

高产量扩增

TopTaq DNA Polymerase配合新型的TopTaq PCR Buffer使用,有利于通过提供高比率的特异性-非特异性引物结合来扩增特异性PCR产物。由于KCl和(NH4)2SO4独特的均衡结合,比传统的PCR缓冲液,该PCR缓冲液提供广泛的退火温度和Mg2+浓度条件以及严格的引物-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过程的需要,有时不需要优化。因此,TopTaq DNA Polymerase提供与Taq DNA Polymerase相同的可靠性和特异性,并且能额外地在一些PCR反应体系中提供优化的性能,得到更多的PCR产物(参见"High yields of PCR product without optimization")。

TopTaq稳定剂

TopTaq稳定剂与TopTaq DNA Polymerase在4°C以及室温下结合,防止聚合酶在长期储存中降解,同时避免模板DNA和引物在低温下与聚合酶结合。在初始的降解步骤中,TopTaq稳定剂从聚合酶中分离,而不影响聚合酶的活性(参见"Stabilization at 4°C")。

CoralLoad Concentrate

为了更方便使用,成套提供TopTaq DNA Polymerase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")。

优化的实验方案和缓冲液

独特的PCR缓冲液,具有均衡的钾和铵离子,配合TopTaq DNA Polymerase使用能促进特异性引物-模板退火,同时减少非特异性退火。即使使用少量模板也能获得最大量的特异性产物。独特的缓冲液制备和单独的预优化实验方案消除了个体-引物体系实验参数优化的需求,节省了时间和成本。甚至使用相同的Mg2+浓度和退火温度扩增大量不同大小的片段,也可获得高产量的特异性扩增产物(参见"High PCR product yields without optimization")。

Q-Solution

新型的PCR辅助剂有助于通过修改DNA熔化行为来扩增困难模板。使用该独特的试剂通常能进行或促进欠佳的PCR反应。不同于DMSO和其他的PCR辅助剂, Q-Solution可在各种引物-模板体系的确定工作浓度下使用,且无毒。

Procedure

由于酶保存液中含有独特的专有TopTaq稳定剂,TopTaq DNA Polymerase是第一款可在4°C下常规储存的试剂盒。在初始的降解步骤中,TopTaq稳定剂可从聚合酶中分离而不影响聚合酶活性。这提高了便利性,能够立即进行反应设置,无需使用冰,节省了解冻过程的时间(参见"TopTaq procedure")。所有组件都可以在室温下相结合,无需冰上工作。独特的缓冲液制备和单独的预优化实验方案消除了单个引物-模板体系实验参数的优化需求。

Applications

TopTaq DNA Polymerase高度适用于所有终点法PCR应用。

Feature
Specifications
Applications PCR, RT-PCR, DNA fingerprinting
dNTP's included No
Enzyme activity 5' -> 3' exonuclease activity
Mastermix No
Reaction type PCR amplification
Real-time or endpoint Endpoint
Sample/target type Genomic DNA and cDNA
Single or multiplex Single
Storage conditions Fridge storage and room temperature setup
With/without hotstart Without Hotstart

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Kit Handbooks
1
For standard and specialized end-point PCR applications without the need for optimization
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