• Optimized for Real Time PCR with TaqMan^® probe
• Easy Preparation
• Increased Specificity

Description

Premix Ex Taq™ (Perfect Real Time) is a 2× convenient premix reagent, specially designed for real time PCR by using TaqMan^®*1 probe. This product combines the high performance of TaKaRa Ex Taq™ HS, which is an enzyme for hot start PCR utilizing Taq antibody, with a newly developed buffer which provides superior specificity, increased amplification efficiency and high aptitude for high-speed real time PCR. Accordingly, a successful real time PCR is promised with high sensitivity, wide dynamic range and accurate quantification. This is supplied with ROX™^*2 Reference Dyes to nomalize the fluorescent signal between reactions, for instruments that are equipped with this option.

[Applicable real time PCR instruments]

• Thermal Cycler Dice™ Real Time System (TaKaRa Cat.#TP800)
• ABI PRISM^® 7000/7700/7900HT,^*3
  Applied Biosystems 7300/7500 Real-Time PCR System
  7500 Fast Real-Time PCR System (Applied Biosystems)
• LightCycler^® (Roche Diagnostics)^*4
• Smart Cycler^® (Cepheid)^*5

*1 TaqMan^® is a registered trademark of Roche Molecular Systems Inc.
*2 ROX™ is a trademark of Applera Corporation or its subsidiaries in the US and/or certain other countries.
*3 ABI PRISM^® is a registered trademark of Applera Corporation.
*4 LightCycler^® is a registered trademark of Roche Diagnostics.
*5 Smart Cycler^® is a registered trademark of Cepheid.

Reagents and Instruments required but not included in the kit

1. Thermal Cycler for real time PCR
2. Reaction tube or plate for real time PCR
3. PCR primers
4. TaqMan^® probe for detection (licensed probes)
5. dH₂O
6. Micropippets and Micropippet tips (autoclaved prior to use)

Storage

Stable at 4°C for 6 months.
Careful attention should be made not to cause contamination.

* This product is shipped at -20°C. After delivered, store at 4°C. Make the concentration uniform by gently inverting a tube before use.
* This product can be stored at -20°C for long term storage. Once thawed, store at 4°C and use up within 6 months.

Content (250 units)

Premix Ex Taq™ (Perfect Real Time) (2×conc) *		1.0 ml × 5 tubes
ROX™ Reference Dye (50×conc.) **		200 μl × 1 tube
ROX™ Reference Dye II(50 × conc.) **		200 μl × 1 tube

* contains TaKaRa Ex Taq ™ HS, dNTP Mixture, Mg²⁺.
** ROX™ Reference Dye/Dye II is supplied for performing normalization of fluorescent signal intensities among wells when used with real time PCR instruments that have this option.

Specification of TaKaRa Ex Taq™ HS

Unit definition: One unit is the amount of the enzyme that will incorporate10 nmol of dNTP into acid-insoluble products in 30 minutes at 74°C with activated salmon sperm DNA as the template-primer.

Reaction mixture for unit definition:

25 mM		TAPS (pH9.3 at 25°C)
50 mM		KCl
2 mM		MgCl2
1 mM		2-mercaptoethanol
200 μM		each dATP, dGTP, dTTP
100 μM		[α-32P]-dCTP
0.25 mg/ml		activated salmon sperm DNA

Purity: Nicking activity, endonuclease and exonuclease activity were not detected after the incubation of 0.6 μg of supercoiled pBR322 DNA, 0.6 μg of λ DNA or 0.6 μg of λ -Hind III digest with 10 units of this enzyme for 1 hour at 74°C.

Application

For real time quantification of DNA using TaqMan^® probe.

Features

Quick and accurate detection and quantification of target gene through real time PCR.

High amplification efficiency and highly sensitive detection:
This kit utilizes an enzyme for hot start, TaKaRa Ex Taq™ HS. As this enzyme-buffer system is optimized for real time PCR, this product offers highly effcient amplification and high sensitivity.

Convenient:
A separate tube of ROX™ reference dye is supplied.

Versatility:
Compatible with Smart Cycler^®, LightCycler^®, ABI PRISM^® 7000/7700/7900HT and 7300/7500/7500 Fast Real-Time PCR System.

Note

TaKaRa Ex Taq™ HS used in this product is an enzyme for hot start PCR utilizing Taq antibody.
Initial denaturation step prior to PCR should be 95°C for 30 sec. It does not require the activation step of heating at 95°C for 10-15 min prior to the PCR.

PCR products:

As most PCR products amplified with TaKaRa Ex Taq™ HS have one A added at 3′-termini, the obtained PCR product can be directly used for cloning into T-vector. Also it is possible to clone the product in blunt-end vectors after blunting and phosphorylation of the end.