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Qiagen/QuantiTect Probe PCR Kits/For 200 x 50 µl reactions: 3 x 1.7 ml 2x QuantiTect Probe PCR Master Mix, 2 x 2 ml RNase-Free Water/204343
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Qiagen/QuantiTect Probe PCR Kits/For 200 x 50 µl reactions: 3 x 1.7 ml 2x QuantiTect Probe PCR Master Mix, 2 x 2 ml RNase-Free Water/204343
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Qiagen
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204343
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QuantiTect Probe PCR Kits

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For real-time PCR and two-step RT-PCR using sequence-specific probes

  • Highly sensitive detection of low-copy targets
  • Accurate quantification over several logs of template
  • Use of any sequence-specific probe on any real-time cycler
  • Available with or without uracil-N-glycosylase (UNG)
  • No need to optimize reaction and cycling conditions

QuantiTect Probe PCR Kits enable sensitive quantification of gDNA and cDNA targets by real-time PCR and two-step RT-PCR using sequence-specific probes. The combination of a hot start and a unique PCR buffer system in the ready-to-use master mix ensures highly sensitive qPCR on any real-time cycler without the need for optimization. The dNTP mix includes dUTP, allowing optional treatment with UNG. For convenience, the master mix can be stored at 2–8°C.

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Cat No./ID:204343
QuantiTect Probe PCR Kit (200)
$479.00
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For 200 x 50 µl reactions: 3 x 1.7 ml 2x QuantiTect Probe PCR Master Mix, 2 x 2 ml RNase-Free Water
Cat No./ID:204345
QuantiTect Probe PCR Kit (1000)
$1,976.00
Order Product
For 1000 x 50 µl reactions: 25 ml 2x QuantiTect Probe PCR Master Mix, 20 ml RNase-Free Water
Cat No./ID:204363
QuantiTect Probe PCR +UNG Kit (200)
$581.00
Order Product
For 200 x 50 µl reactions: 3 x 1.7 ml 2x QuantiTect Probe PCR Master Mix, 100 ul UNG Solution, 2 x 2 ml RNase-Free Water
QuantiTect Probe PCR Kits are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

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Product Details

9
Wide dynamic range in two-step PCR.

Duplicate reactions were performed on the Mx3005P using tenfold dilutions of human keukocyte cDNA (10 ng to 0.01 ng) and a TaqMan assay for IL1R2 (a cytokine). The QuantiTect Probe PCR Kit provided accurate gene expression analysis from low to high template amounts with a PCR efficiency of 101%.

9
High sensitivity and efficiency, and wide dynamic range.
Tenfold serial dilutions of leukocyte cDNA (100 ng to 1 pg) were analyzed in duplicate on the Mx3005 using the QuantiTect Probe PCR Kit with primers and a FAM-labeled probe specific for IL8 (interleukin 8). A PCR efficiency of 97% over 6 logs of template dilution was achieved, and as little as 1 pg of IL8 transcript was sensitively detected.
9
Specific primer annealing.
9
Highly specific amplification.
9
Wide dynamic range in real-time PCR.
Probe-based real-time PCR with UNG pretreatment was carried out using the QuantiTect Probe PCR +UNG Kit. Reactions were run in duplicate on the ABI PRISM 7900 using 10-fold dilutions of human leukocyte cDNA (100 ng to 10 pg) and a TaqMan assay for IL8 (a cytokine). The amplification plots were evenly spaced, leading to a high PCR efficiency of 94%.
9
Two-step RT-PCR.
9
Effective UNG digestion.
106 copies of two dUMP-containing PCR amplicons were treated with or without UNG and then amplified by real-time PCR. UNG was from various suppliers, and real-time PCR was performed using the master mix from the QuantiTect Probe PCR +UNG Kit. ΔCT on the Y-axis indicates CT values for non-UNG-treated samples subtracted from CT values for UNG-treated samples. The UNG supplied with the QuantiTect Kit provided the greatest ΔCT (9-12 cycles). This indicates that QIAGEN UNG digests carryover PCR products more effectively than UNG from other suppliers.
Performance

The QuantiTect Probe PCR Kit delivers sensitive and reliable results in combination with the QuantiTect Reverse Transcription Kit (see figure "High sensitivity and efficiency, and wide dynamic range"). The unique PCR buffer composition enables QuantiTect Probe PCR Kits to provide sensitive quantification of low-copy DNA targets, as well as accurate quantification over a wide linear range (see figures "High sensitivity and efficiency, and wide dynamic range" and "Wide dynamic range in two-step RT-PCR"). HotStarTaq DNA Polymerase further increases the specificity of the PCR reaction by providing the most stringent hot start compared with other polymerases (see figure "Highly specific amplification").

A major source of PCR contamination is the carryover of PCR products from previous reactions. Pretreatment with UNG prior to starting PCR ensures that any contaminating PCR products do not affect subsequent PCR. The combination of the specially optimized UNG solution and the proven PCR master mix in the QuantiTect Probe PCR +UNG Kit ensures effective elimination of carried-over PCR products together with reliable quantification of target sequences (see figures "Effective UNG digestion" and "Wide dynamic range in real-time PCR").

Principle

QuantiTect Probe PCR Kits contain an optimized, ready-to-use master mix for highly specific and sensitive real-time quantification of gDNA and cDNA targets using sequence-specific probes.The kits are designed for use with all types of sequence-specific probes, including hydrolysis probe detection (e.g., TaqMan® and other dual-labeled probes), FRET probes, and Molecular Beacons. QuantiTect Probe PCR Kits containa unique PCR buffer that containsa balanced combination of K+ and NH4+ ions, which promote specific primer annealing, enabling high PCR specificity and sensitivity (see figure "Specific primer annealing"). In addition, HotStarTaq DNA Polymerase provides a stringent hot start, preventingthe formation of nonspecific products.

QuantiTect Probe PCR Master Mix also contains dUTP, enabling pretreatment with uracil-N-glycosylase (UNG) prior to starting PCR, which ensures that any contaminating PCR products do not affect subsequent PCR reactions.

Components of 2x QuantiTect Probe PCR Kit
Component Features Benefits
HotStarTaq DNA Polymerase 15 minactivation at 95ºC Set-up of qPCR reactions at room temperature
QuantiTect Probe PCR Buffer Balanced combination of NH4+ and K+ ions Specific primer annealing ensures reliable PCR results
dNTP mix Includes dUTP, which partially replaces dTTP and enables optional UNG treatment of reactions Eliminatescontamination from carryover of PCR products by optional UNG treatment
ROX dye For normalization of fluorescent signals on Applied Biosystems and, optionally, Agilent instruments Precise quantification on cyclers that require ROX dye.Does not interfere with reactions on other real-time cyclers
Procedure

QuantiTect Probe PCR Kits overcome the need for optimization of reaction conditions, which can be tedious and time-consuming. Simply add primers, probe, and DNA template to the ready-to-use PCR master mix, and start the reaction (see flowchart "Two-step RT-PCR"). Follow the protocol in the handbook to get fast and reliable results on any real-time cycler. If required, reactions can be pretreated with uracil-N-glycosylase (UNG) to eliminate carryover of PCR products from previous reactions. 

For optimal results in real-time two-step RT-PCR, we recommend synthesizing cDNA using the QuantiTect Reverse Transcription Kit. The kit provides fast cDNA synthesis in just 20 minutes with integrated removal of genomic DNA contamination.

Applications

QuantiTect Probe PCR Kits can be used for probe-based gene expression analysis of cDNA targets or quantitative gDNA analysis on any real-time cycler. This includes instruments from Applied Biosystems, Bio-Rad, Cepheid, Eppendorf, Roche, and Agilent. For the Rotor-Gene Q and other Rotor-Gene cyclers, we recommend using the Rotor-Gene Probe PCR Kit, which has been specially developed for fast cycling on these instruments.

Specifications

Features
Specifications
ApplicationsReal-time quantification of DNA, cDNA, or RNA targets
Reaction typePCR and two-step RT-PCR
Real-time or endpointReal-time
Sample/target typeDNA, cDNA, RNA
Single or multiplexSingle
SYBR Green I or sequence-specific probesSequence-specific probes
Thermal cyclerMost real-time cyclers (e.g. LC, RG, ABI)
With or without ROXWith ROX

Product Resources

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Brochures & Guides (2)
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(EN) - Analyzing Gene Expression and Regulation
Show details
Critical Factors for Successful Real-Time PCR - (EN)
Show details
FAQs (30)
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Do you recommend 1-step or 2-step real-time RT-PCR for gene expression analysis?
FAQ ID -1056
View
What should I use as a standard for absolute quantification in real-time PCR?
FAQ ID -1085
View
How much template can I use in the reaction and what is the maximum volume of template that can be used in the QuantiTect Kits?
FAQ ID -1086
View
How much time will be saved when switching from standard cycling to fast cycling with QuantiFast Kits?
FAQ ID -1438
View
How do QuantiFast PCR Kits compare to QuantiTect PCR Kits for quantitative real-time PCR?
FAQ ID -1441
View
Why is the storage time for QuantiFast PCR Kits shorter than that for QuantiTect PCR Kits?
FAQ ID -1446
View
Why is the reaction volume for QuantiFast PCR Kits lower than that for QuantiTect PCR Kits?
FAQ ID -1447
View
Which real-time PCR kits are recommended downstream of the QuantiTect Whole Transcriptome Kit?
FAQ ID -1592
View
Do the master mixes in Rotor-Gene Kits contain dUTP to allow UNG pretreatment?
FAQ ID -2117
View
What are the main differences between Rotor-Gene and QuantiTect or QuantiFast PCR Kits?
FAQ ID -2119
View
Can Uracil-N-Glycosylase (UNG) be purchased separately, independent of QuantiTect PCR Kits?
FAQ ID -2134
View
Are QuantiTect PCR +UNG Kits available in bulk format (e.g., 25 ml master mix)?
FAQ ID -2136
View
Can Uracil-N-Glycosylase (UNG) be added to the QuantiTect PCR Master Mix and be stored for later use?
FAQ ID -2138
View
Will Uracil-N-Glycosylase (UNG) completely remove contaminating amplicons when using QuantiTect +UNG Kits?
FAQ ID -2139
View
I would like to order the QuantiFast Probe Assays without master mix. Is this possible?
FAQ ID -2360
View
Do you have any information or guidelines regarding the choice of reference genes for real-time PCR?
FAQ ID -2371
View
What is the threshold cycle or Ct value?
FAQ ID -2682
View
My QuantiTect Master Mix did not freeze at degrees. Is it still ok to use?
-20
View
Why do replicates in real-time PCR have different plateau heights?
FAQ ID -539
View
Do I need to calibrate my real-time cycler if I want to use Yakima Yellow?
FAQ ID -540
View
What is a quenched FRET assay?
FAQ ID -542
View
How many template copies can I detect using QuantiTect Kits in real-time PCR?
FAQ ID -543
View
What reaction volume should I use with QuantiTect Kits?
FAQ ID -551
View
Is it possible to perform a UNG treatment when using QuantiTect kits?
FAQ ID -564
View
Can I make a master mix with QuantiProbes, primers, and all other reaction components and store it for later use?
FAQ ID -584
View
Why are my realtime PCR amplification plots hook-shaped?
FAQ ID -587
View
Why does my realtime PCR assay quality decrease over time?
FAQ ID -589
View
Do you have information on the use of recombinant DNA and RNA as absolute standards for realtime RT-PCR?
FAQ ID -729
View
Why should DNA or cDNA targets be less than 250 bp long for real-time PCR?
FAQ ID-751
View
How should fluorescent labeled probes be stored?
FAQ ID -784
View
Kit Handbooks (2)
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Uracil-N-glycosylase - (EN)
For use with QuantiTect PCR Kits to eliminate carryover of PCR products
Show details
(EN) - QuantiTect Probe PCR Handbook
For quantitative, real-time PCR and two-step RT-PCR using sequence-specific probes
Show details
Technical Information (1)
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(EN) - Critical factors for synthesis of cDNA for real-time PCR analysis
Show details
Scientific Posters (1)
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(EN) - Workflow for DNA purification from tough specimens
Show details
Quick-Start Protocols (1)
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QuantiTect Probe PCR Quick-Start Protocol
Show details
Safety Data Sheets (4)
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MSDS QuantiTect Probe PCR Kit (200)
MSDS QuantiTect Probe PCR Kit (1000)
MSDS QuantiTect Probe PCR +UNG Kit (200)
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