HotStarTaq Plus DNA Polymerase

별도로 조건을 setup할 필요없이 특이성이 높은 hot-start PCR을 수행할 수 있습니다

이 제품은 2023년 11월 30일에 단종됩니다.

지금 바로 후속 AllTaq PCR 키트로 바꿔 보세요. 전환에 대해 자세히 알아보고 상세한 FAQ를 확인해 보세요.

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HotStarTaq Plus DNA Polymerase (1000)

Cat. No. / ID: 203605

1000 units HotStarTaq Plus DNA Polymerase, 10x PCR Buffer, 10x CoralLoad PCR Buffer, 5x Q-Solution, 25 mM MgCl₂

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HotStarTaq Plus DNA Polymerase은/는 분자생물학 분야에 사용하기 위한 것입니다. 이 제품은 질병의 진단, 예방, 또는 치료용이 아닙니다.

이 제품은 2023년 11월 30일에 단종됩니다.

지금 바로 후속 AllTaq PCR 키트로 바꿔 보세요. 전환에 대해 자세히 알아보고 상세한 FAQ를 확인해 보세요.

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특징

High PCR specificity — 뛰어한 HotStarTaq DNA Polymerase의 PCR 수행 능력
빠른 5분 enzyme activiation 시간과 최소한의 optimization — 시간 절약, 경제적
빠르고 편리함을 제공하는 PCR loading buffer가 들어 있음

제품 세부 정보

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.

성능

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 ""). Together, these components ensure specific amplification in a range of applications (see figure "Effect of hot start on RT-PCR performance" and "").

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: ≥10⁵ 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

그림 참조

Amplification of difficult templates.

Highly sensitive single-cell PCR.

원리

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 (NH₄)₂SO₄, the buffer provides stringent primer-annealing conditions over a wider range of annealing temperatures and Mg²⁺ concentrations than conventional PCR buffers. Optimization of PCR by varying the annealing temperature or the Mg²⁺ 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.

그림 참조

Increased specificity of primer annealing.

CoralLoad PCR Buffer.

Amplification of difficult templates.

절차

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.

응용 분야

HotStarTaq Plus DNA Polymerase is highly suitable for a wide variety of applications, including challenging applications such as amplification of:

Complex genomic templates
Complex cDNA templates (e.g., RT-PCR)
Very low-copy targets (e.g., single-cell PCR)
Reactions with multiple primer pairs

지원되는 데이터 및 수치

CoralLoad PCR Buffer.

Effect of hot start on RT-PCR performance.

Higher specificity with different primer–template systems.

Specifications

Features	Specifications
Applications	PCR, RT-PCR, Complex genomic templates, very low-copy targets
With/without hotstart	With hotstart
Sample/target type	Genomic DNA and cDNA
Enzyme activity	5' -> 3' exonuclease activity
Reaction type	PCR amplification
Single or multiplex	Single
Mastermix	No
Real-time or endpoint	Endpoint

리소스

Second edition — innovative tools

Addressing critical factors and new solutions

For highly specific hot-start PCR without optimization

Publications

The putatively functional Mkrn1-p1 pseudogene is neither expressed nor imprinted, nor does it regulate its source gene in trans.

Gray TA; Wilson A; Fortin PJ; Nicholls RD;

Proc Natl Acad Sci U S A; 2006; 103 (32):12039-44 2006 Aug 1 PMID:16882727

Use of a short fragment of the C-terminal E gene for detection and characterization of two new lineages of dengue virus 1 in India.

Domingo C; Palacios G; Jabado O; Reyes N; Niedrig M; Gascón J; Cabrerizo M; Lipkin WI; Tenorio A;

J Clin Microbiol; 2006; 44 (4):1519-29 2006 Apr PMID:16597885

FAQ

No. HotStarTaq Plus DNA Polymerase is supplied with conventional QIAGEN PCR Buffer and CoralLoad PCR Buffer in separate vials. Both buffers minimize nonspecific amplification products, primer–dimers, and background.

CoralLoad PCR Buffer 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.

FAQ ID -1052

PCR products that will be cloned using the QIAGEN PCR Cloning Kit should be generated using a thermostable DNA Polymerase without proofreading activity, such as Taq DNA Polymerase. Such polymerases attach a single A overhang to their reaction products, which can hybridize to the U overhang of the pDrive Cloning Vector. For efficient addition of an A overhang during the PCR procedure, we recommend a final extension step for 10 min at 72°C as described in the standard protocols of the Taq PCR- and HotStarTaq PCR handbook.

FAQ ID -165

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

Yes. Do not increase the activation time for HotStarTaq Plus DNA Polymerase beyond the recommended 5 minutes, as PCR product yield may be reduced and less specific amplification may result. Please follow the instructions in the HotStarTaq Plus PCR Handbook closely, and use the cycling parameters presented in the handbook to ensure optimal results.

FAQ ID -1050

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

The activation time for HotStarTaq Plus DNA Polymerase used in the QuantiFast SYBR Green PCR Kits is longer than that for QuantiFast Probe PCR Kits. This is due to differences in buffer composition. Buffer components such as salts and additives influence the time required for enzyme activation.

FAQ ID -1449

Yes. Separated dyes are visible on an agarose gel down to a volume of 2 µl of PCR product generated with HotStarTaq Plus DNA Polymerase and CoralLoad PCR buffer.

FAQ ID -1051

In QIAGEN labs, we have amplified PCR products up to 5 kb from complex genomic DNA, and up to 10 kb from less complex lambda phage DNA with the HotStar HiFidelity Polymerase Kit, following standard protocols in the HotStar HiFidelity PCR Handbook.

For targets larger than 5 kb of complex genomic DNA, and larger than 10 kb of less complex DNA, we recommend to follow the protocol 'Amplification of Long PCR Products' in the HotStar HiFidelity PCR Handbook. The protocol uses a mixture of HotStar HiFidelity DNA Polymerase and Taq, or HotStar Taq Plus DNA Polymerase, and allows much longer fragments to be generated. In-house we have tested fragments up to 13 kb from complex genomic DNA or up to 30 kb from less complex lambda phage DNA using this protocol.

FAQ ID -1047