RT² SYBR^® Green FAST Mastermixes

For use with 100-ring disc format RT² Profiler and RT² lncRNA PCR Arrays on the Rotor-Gene Q

Products

RT² SYBR^® Green FAST Mastermixes are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

RT² SYBR Green ROX FAST Mastermix (2)

Cat. No. / ID: 330620

Contains 2 x 1.35 ml tubes: for 12 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 250 x 20 µl reactions

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RT² SYBR Green FAST Mastermix (8)

Cat. No. / ID: 330601

Contains 8 x 1.35 ml tubes: for 8 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 1000 x 20 µl reactions

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RT² SYBR Green ROX FAST Mastermix (24)

Cat. No. / ID: 330623

Contains 24 x 1.35 ml tubes: for 24 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 3000 x 20 µl reactions

7 855,00 MYR

Quantity

RT² SYBR Green ROX FAST Mastermix (25 ml)

Cat. No. / ID: 330629

Contains 1 bottle of 25 ml mastermix: for 24 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 2500 x 20 µl reactions

6 840,00 MYR

Quantity

RT² SYBR Green FAST Mastermix (12)

Cat. No. / ID: 330602

Contains 12 x 1.35 ml tubes: for 12 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 1500 x 20 µl reactions

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RT² SYBR Green ROX FAST Mastermix (8)

Cat. No. / ID: 330621

Contains 8 x 1.35 ml tubes: for 8 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 1000 x 20 µl reactions

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RT² SYBR Green FAST Mastermix (2)

Cat. No. / ID: 330600

Contains 2 x 1.35 ml tubes: for 2 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 250 x 20 µl reactions

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RT² SYBR Green ROX FAST Mastermix (12)

Cat. No. / ID: 330622

Contains 12 x 1.35 ml tubes: for 12 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 1500 x 20 µl reactions

Inquire

RT² SYBR Green FAST Mastermix (24)

Cat. No. / ID: 330603

Contains 24 x 1.35 ml tubes: for 24 x 100-well ring discs on the QIAGEN Rotor-Gene Q or 3000 x 20 µl reactions

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Features

Designed for 100-ring disc RT² Profiler / RT² lncRNA PCR Arrays
Available with ROX, fluorescein, or without dye
Well-suited for use with self-designed PCR assays
Exceptional qPCR performance in instruments with fast cycling conditions

Product Details

RT²SYBR Green FAST Mastermixes are specially formulated for use with the 100-ring disc format of the RT² Profiler and RT² lncRNA PCR arrays on the Rotor-Gene Q. This mastermix is suited for use with self-designed qPCR assays on other types of real-time PCR instruments with fast cycling conditions. RT² SYBR Green FAST Mastermixes are available with ROX, fluorescein, or without reference dyes.

Performance

RT² SYBR Green FAST Mastermixes contain all of the optimized reagents and buffers needed for SYBR Green based real-time PCR with RT² qPCR Primer Assays and RT² Profiler PCR Arrays on all real-time PCR instruments.

The high-performance real-time PCR enabled by RT² SYBR Green FAST Mastermixes demonstrates high amplification efficiencies (see figure " High amplification efficiency over a wide dynamic range") and high levels of sensitivity and specificity (see figure " Tighter control of polymerase activity yields greater specificity").

In addition, dissociation curves obtained using RT² SYBR Green FAST Mastermixes have lower background and a sharper shape compared with dissociation curves generated by regular cycling conditions (see figure " Sharper dissociation curves with RT² SYBR Green Fast Mastermix").

See figures

High amplification efficiency over a wide dynamic range.

Tighter control of polymerase activity yields greater specificity.

Sharper dissociation curves with RT2 SYBR^® Green FAST Mastermix.

Principle

RT² SYBR Green FAST Mastermixes contain real-time PCR buffer, a high-performance, HotStart DNA Taq polymerase, nucleotides, and SYBR Green dye. Some mastermixes contain either fluorescein or ROX dye for optimization of the instrument optics. The chemically-modified and tightly controlled HotStart enzyme uniquely provides accurate SYBR Green results by preventing the amplification of primer–dimers and other nonspecific products. RT² SYBR Green FAST Mastermixes are optimized to use a 10-second denaturation step and 30-second annealing/extension step PCR protocol, without compromising specificity or sensitivity.

Mastermixes without ROX or fluorescein

RT² SYBR Green FAST Mastermix is highly suited for real-time PCR applications using SYBR Green-based detection on instrumentation not requiring a reference dye, including Bio-Rad models CFX96, CFX384; Bio-Rad/MJ Research Chromo4, DNA Engine Opticon, DNA Engine Opticon 2; Roche LightCycler 480 (96-well and 384-well); Eppendorf Mastercycler ep realplex without ROX filter set; and Cepheid SmartCycler.

Mastermixes with fluorescein

RT²SYBR Green Fluor FAST Mastermix is highly suited for real-time PCR applications using SYBR Green-based detection on instrumentation that uses fluorescein as a reference dye, including the Bio-Rad models iCycler, iQ5, MyiQ, and MyiQ2.

Mastermixes with ROX

RT² SYBR Green ROX FAST Mastermix is highly suited for real-time PCR applications using SYBR Green-based detection on instrumentation that uses ROX as a reference dye, including QIAGEN’s Rotor-Gene Q; Applied Biosystems models 5700, 7000, 7300, 7500 (Standard and Fast), 7700, 7900HT (Standard and Fast 96-well block, 384-well block), StepOnePlus, ViiA 7 (Standard and Fast 96-well block, 384-well block); Eppendorf Mastercycler ep realplex with or without ROX filter set; Stratagene models Mx3000P, Mx3005P, Mx4000; and Takara TP-800.

Applications

RT² SYBR Green FAST Mastermixes are suitable for use in real-time PCR applications.

Supporting data and figures

Sharper dissociation curves with RT2 SYBR^® Green FAST Mastermix.

A positive PCR control was amplified on the ABI 7500 qRT-PCR instrument using [A] regular cycling and RT² SYBR Green Mastermix or [B] fast cycling with RT² SYBR Green FAST Mastermix. The dissociation curve obtained with RT² SYBR Green FAST Mastermix has lower background and a sharper shape.

Resources

For pathway-focused gene expression profiling using real-time RT-PCR

Download Safety Data Sheets for QIAGEN product components.

FAQ

There are less than a dozen genes encoded by the mitochondrial genome (all other mitochondrial proteins are encoded by nuclear genes), and they are all transcribed as one transcript (just like any prokaryote), so distinguishing the expression of individual genes by real-time RT-PCR is not possible.

FAQ ID -2680

Be sure to carefully and completely seal the qPCR assay plate with fresh, optical, thin-wall, 8-cap strips or adhesive optical film before the plate is placed into the real-time cycler. In addition, refer to your instrument's user's manual to determine whether the real-time cycler manufacturer recommends use of a plate compression pad during the run.

FAQ ID -2679

Any DNA contamination will artificially inflate the SYBR Green signal, yielding skewed gene expression profiles and false-positive signals. The most common source of DNA contamination is from PCR products generated during previous experiments. Such contamination is most often due to the improper disposal of tubes, tips, and gels that previously came into contact with PCR products. Additionally, PCR products may also contaminate pipettors, racks, work pads, and commonly used reagents such as water and buffers. To minimize the risk of contaminating your experiment with extraneous DNA, the following steps should be taken:

Remove a single aliquot of water from your PCR-grade stock, sufficient to complete the experiment. This minimizes the number of times that the stock container is opened, thereby minimizing contamination risks.
Use only fresh PCR-grade reagents and disposable labware.
Treat any labware (tubes, tips, and tip boxes) used in PCR with 10% bleach, before discarding.
Maintain a dedicated workspace for PCR setup (perhaps a PCR-only hood), away from areas of the lab where post-PCR work is done, such as running gels, enzyme digestions, and cloning.
Change the lab bench pads/papers often and decontaminate lab benches and labware (racks, pipettors, etc.) before each use by washing with 10% bleach, and/or exposing to UV light for at least 10 minutes. This serves to degrade and/or inactivate contaminating DNA.
Before, during, and after the experiment, minimize the opening and closing of any tubes or plates used during the experiment.

FAQ ID -2654

Dissociation curves are carried out at the end of a PCR experiment by following a 3-step procedure.

First, all the components are denatured at 95°C, followed by complete annealing at a set temperature (based on the primer Tm values), followed by a gradual increase in temperature up to 95°C. Fluorescence intensity is monitored during this final temperature increase, resulting in the generation of a melting curve or dissociation curve.

By analyzing the first derivative of such a curve, you can readily assess the homogeneity of the PCR products, including the presence of primer–dimers, thereby determining the specificity of the PCR reaction. It is important to carry out such post-PCR analyses when using SYBR Green probe chemistry due to this reagent's lack of sequence specificity.

FAQ ID -2678

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