Rotor-Gene Multiplex PCR Kit

For ultrafast, multiplex, real-time PCR and two-step RT-PCR on Rotor-Gene cyclers


  • Optimized for ultrafast, reliable results on Rotor-Gene cyclers
  • Sensitive detection of up to 4 targets in 1 tube
  • Successful multiplex PCR without the need for optimization
  • Reliable quantification of low- and high-abundance targets

Product Details

The Rotor-Gene Multiplex PCR Kit is designed for use with the Rotor-Gene Q and other Rotor-Gene cyclers, providing ultrafast, highly reliable quantification in multiplex, real-time PCR and two-step RT-PCR using sequence-specific probes. Depending on the cycler configuration, up to 4 cDNA or gDNA targets (e.g., 1 control gene and 3 target genes) can be quantified simultaneously in the same tube. Outstanding performance is achieved through the combination of a specially optimized master mix and the unique Rotor-Gene cycler. For convenience, the master mix can be stored at 2–8°C.


IMPORTANT NOTE: As announced earlier, the production of the Rotor-Gene kits has been discontinued since mid-2021. Hence, these products will be available only until stocks last. Visit the product page of the successor kit to view improved features or to request a trial kit.


For more information and FAQs on this transition, visit:


With the Rotor-Gene Multiplex PCR Kit, up to 4 cDNA targets can be simultaneously and rapidly quantified in the same tube, increasing throughput and saving precious sample material (see figures " Reliable multiplex analysis" and " Reliable duplex analysis"). Genes of different expression levels are all amplified in the same tube with the same high efficiency, enabling reliable relative quantification of gene expression.
See figures


Amplifying reference and target genes in the same reaction instead of in separate reactions increases the reliability of gene quantification by minimizing handling errors. The Rotor-Gene Multiplex PCR Kit enables reliable multiplex real-time PCR quantification of cDNA targets on the Rotor-Gene Q without the need for optimization of reaction and cycling conditions (see flowchart " QIAGEN multiplex kits"). Highly specific amplification is assured through a balanced combination of K+ and NH4+ ions, which promote specific primer annealing and enable high PCR specificity and sensitivity, while synthetic Factor MP, an innovative PCR additive specially developed for challenging multiplex PCR applications, allows different amplicons in the same reaction to all be amplified with the same high efficiency (see figure " Unique PCR buffer").

Fast cycling without compromising performance is achieved using Q-Bond, a novel PCR additive that considerably shortens cycler run times (see figure " Fast primer annealing"). In addition, the highly stringent hot-start enzyme HotStarTaq Plus DNA Polymerase is rapidly activated at the start of PCR by a brief 5-minute incubation at 95ºC. 

Components of 2x Rotor-Gene Multiplex PCR Kit*
ComponentFeatures Benefits
HotStarTaq Plus DNA Polymerase 5 min activation at 95ºC Set up of qPCR reactions at room temperature
Rotor-Gene Multiplex PCR Buffer Balanced combination of NH4+ and K+ ions Specific primer annealing ensures reliable qPCR results
Synthetic Factor MP Reliable multiplexing analysis of up to 4 targets in the same tube
Unique Q-Bond additive Faster PCR run times enable faster results and more reactions per day
* Also contains dNTP mix (dATP, dCTP, dGTP, dTTP).
See figures


A ready-to-use master mix eliminates the need for optimization of reaction and cycling conditions. Simply add template cDNA and primer-probe sets to the master mix and program the cycler. The handbook supplied with the kit lists recommended dyes and contains a single protocol for all multiplex qPCR assays.

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.


The Rotor-Gene Multiplex PCR Kit is optimized for fast, real-time PCR and two-step RT-PCR analysis using sequence-specific probes on the Rotor-Gene Q. It is also compatible with the Rotor-Gene 3000 and the Rotor-Gene 6000. Up to 4 cDNA or gDNA targets can be simultaneously and rapidly quantified in the same tube, increasing throughput and saving precious sample material.

For ultrafast, one-step qRT-PCR multiplex analysis of RNA targets using sequence-specific probes on Rotor-Gene cyclers, use the Rotor-Gene Multiplex RT-PCR Kit.

Supporting data and figures


ApplicationsReal-time quantification of genomic DNA or cDNA targets in a multiplex format
Real-time or endpointReal-time
Reaction typeReal-time PCR and two-step RT-PCR
Sample/target typeDNA, cDNA
DescriptionFor ultrafast quantitative multiplex real-time PCR and two-step RT-PCR using sequence-specific probes
Single or multiplexMultiplex
SYBR Green I or sequence-specific probesSequence-specific probes
Thermal cyclerRotor-Gene Q, Rotor-Gene 3000, Rotor-Gene 6000
With or without ROXWithout ROX dye


Safety Data Sheets (1)
Download Safety Data Sheets for QIAGEN product components.
Brochures & Guides (1)
Now with even more applications!
Kit Handbooks (1)
For fast multiplex real-time PCR, two-step RT-PCR, and one-step RT-PCR using sequence-specific probes on Rotor-Gene cyclers


Do you have any information or guidelines regarding the choice of reference genes for real-time PCR?

Yes, please visit our website section 'Using endogenous control genes in real-time RT-PCR' for general information. It provides a list of relative gene expression levels for commonly used human and mouse reference genes.

We offer a set of ready-to-order control genes for use in SYBR Green based as well as probe based real-time RT-PCR.

In addition, you may want to refer to the following citations on reference gene selection for quantitative real-time PCR:

• Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, DePaepe A, Speleman F [2002]: Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 2002, 3:0034.

• Radonic A, Thulke S, Mackay IM, Landt O, Siegert W, Nitsche A., 2004. Guideline to reference gene selection for quantitative real-time PCR. Biochem Biophys Res Commun. 313(4): 856-62.

• Katrien Smits,Karen Goossens, Ann Van Soom, Jan Govaere, Maarten Hoogewijs, Emilie Vanhaesebrouck,Cesare Galli, Silvia Colleoni, Jo Vandesompele, and Luc Peelman [2009]Selection of reference genes for quantitative real-time PCR in equine in vivo and fresh and frozen-thawed in vitro blastocysts. BMC Res Notes. Dec 11;2:246.

FAQ ID -2371
Do the master mixes in Rotor-Gene Kits contain dUTP to allow UNG pretreatment?

No. The master mixes in Rotor-Gene Kits contain dTTP instead of dUTP. If UNG treatment is required, we recommend using QuantiTect +UNG Kits. QuantiTect Kits are also compatible with the Rotor-Gene Q; however, the kits require a significantly longer cycling time.



FAQ ID -2117
If housekeeping and target genes differ significantly in abundance, will they both be amplified with equal efficiency using the Rotor-Gene Multiplex PCR Kit?

Yes. The optimized master mix of the Rotor-Gene Multiplex PCR Kit ensures that all gene targets in a multiplex reaction are amplified with the same efficiency and sensitivity as in corresponding singleplex reactions, independent of starting copy numbers.


FAQ ID -2129
Are probes other than TaqMan® probes, such as FRET probes, compatible with fast cycling with the Rotor-Gene Multiplex PCR Kit?

The Rotor-Gene Multiplex PCR Kit has been developed primarily for use with TaqMan® probes. Protocols for probes other than TaqMan® probes have not been tested.

See trademarks

FAQ ID -2130
Why is the initial activation step different for Rotor-Gene Probe, SYBR Green and Multiplex Kits?

The buffer composition, which affects the initial reactivation of HotStarTaq Plus DNA Polymerase, has been optimized for each respective Rotor-Gene Kit.


FAQ ID -2118
How do you achieve fast cycling, yet still deliver the same performance in PCR as that achieved with standard cycling?

Our unique multiplex PCR buffer system with ammonium and potassium ions and Factor MP has been further optimized in QuantiFast and Rotor-Gene Kits. We have also discovered Q-Bond, a buffer component which supports the rapid formation of the polymerase–primer–template complex, leading to reduced annealing times.

FAQ ID -1430
Do limiting primer concentrations need to be determined when using the Rotor-Gene Multiplex PCR Kit?

No, that is not necessary. Simply use the primer concentrations specified in the protocols in the handbook supplied with your Rotor-Gene Multiplex PCR Kit.



FAQ ID -2128
What real-time cycler should I use for my qPCR experiments?
There are several manufacturers of high-quality real-time cyclers. These include QIAGEN's Rotor-Gene Q, Applied Biosystems, BioRad, Stratagene, Eppendorf, Roche, TaKaRa, Fluidigm, and Cepheid. The important thing to keep in mind is that, once you select an instrument to use, you must use compatible Rotor-Gene Discs and tubes, 96 or 384 well plates, and qPCR master mixes that are optimized for use in that particular instrument.  For example, QIAGEN's Rotor-Gene SYBR Green, Probe, and Multiplex real-time master mixes.  
FAQ ID -2670
Are Rotor-Gene Kits compatible with reaction setup using the QIAgility instrument?

The majority of the Rotor-Gene Kit data shown in our literature has been generated with the help of the QIAgility instrument. We did not observe any problems during the pipetting steps.


FAQ ID -2116
Are TaqMan® Gene Expression Assays from Applied Biosystems compatible with the Rotor-Gene Multiplex PCR Kit?

Yes, simply use the assays at a final concentration of 1x with the Rotor-Gene Multiplex PCR Kit.

See trademarks

FAQ ID -2131
What are the main differences between Rotor-Gene and QuantiTect or QuantiFast PCR Kits?

Rotor-Gene Kits are specifically developed for the Rotor-Gene Q PCR Cycler. The unique rotary system of the cycler combined with the kits’ proprietary buffer system enable ultrafast cycling. Rotor-Gene Kits do not contain ROX dye since no normalization to a passive reference is required. Also, Rotor-Gene Kits do not contain dUTP; therefore, UNG pretreatment is not possible.


FAQ ID -2119
Will Rotor-Gene Kits also work on the Rotor-Gene 6000 and 3000 cyclers?

Yes. Rotor-Gene Kits will also work on the Rotor-Gene 6000 and Rotor-Gene 3000 PCR cyclers with the cycling conditions specified in the Rotor-Gene kit handbooks.


FAQ ID -2121