For ultrafast, real-time PCR and two-step RT-PCR using sequence-specific probes
- Sensitive detection of even low copy numbers
- Accurate detection of a wide range of template amounts
- Optimized for ultrafast, reliable results on Rotor-Gene cyclers
- Specially formulated, ready-to-use master mix for fast cycling
The Rotor-Gene Probe PCR Kit is designed for use with the Rotor-Gene Q and other Rotor-Gene cyclers, providing ultrafast, highly sensitive quantification of gDNA and cDNA targets with real-time PCR and two-step RT-PCR using sequence-specific probes. 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.
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Rotor-Gene Probe PCR Kit (400)
For 400 x 25 µl reactions: 3 x 1.7 ml 2x Rotor-Gene Probe PCR Master Mix, 2 x 2 ml RNase-Free Water
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204374
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Rotor-Gene Probe PCR Kit适用于分子生物学应用。该产品不适用于疾病的诊断、预防或治疗。
See trademarks.
灵敏、可重复的检测。|real-time PCR准确度高。|对1个细胞也能可靠检测。|宽泛的线性范围。|Specific primer annealing.|Fast primer annealing.|
使用2倍稀释的基因组DNA（30 ng到3.75 ng）和自行设计的TaqMan探针对IL1R2（白介素1受体Ⅱ）进行real-time PCR。反应使用[A] Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR仪或[B]供应商AII提供的试剂盒和循环仪（每个稀释样本重复五次反应）。Rotor-Gene体系能够检测到较低的CT值，有更好的重复性以及对不同浓度模板的较高的分辨率。|使用Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR分析仪对人基因组DNA进行real-time PCR分析。[A] 2倍稀释的DNA（从30 [5000拷贝]到0.06 ng[20拷贝]）使用自行设计的TaqMan探针对IL1R2（白介素1受体Ⅱ）进行分析；每个浓度模板重复五次。相邻浓度模板间的CT值平均差1.07个循环。[B] 5 ng DNA使用自行设计的TaqMan探针对IL1R2进行分析。100次重复实验的标准偏差为0.21。[C] 20 ng DNA使用自行设计的TaqMan探针对ACTB（β-actin）进行分析。100次重复实验的标准偏差为0.29。|使用4倍稀释的人基因组DNA（相当于从16, 666个细胞到1个细胞）和自行设计的TaqMan探针对ACTB（β-actin）进行real-time PCR分析。反应使用Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR分析仪进行，重复三次实验。Rotor-Gene体系在整个浓度范围内检测可靠且重复性好。|使用10倍稀释的质粒DNA（109 到10个拷贝）和自行设计的TaqMan探针对PPIA（亲环素A）进行real-time PCR分析。反应使用Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR分析仪进行，重复三次实验。Rotor-Gene体系在整个浓度范围内检测可靠且CT值重复性好。|Cations in the Rotor-Gene Q PCR buffer increase specific primer annealing. K+ binds to phosphate groups on double-stranded DNA, stabilizing primer annealing. NH4+ destabilizes weak hydrogen bonds between mismatched bases.|[A] Q-Bond in Rotor-Gene Probe PCR Master Mix increases the affinity of DNA polymerase for short single-stranded DNA, reducing primer annealing time to a few seconds. In addition, the unique buffer composition supports the melting of DNA, reducing denaturation and extension times. [B] Without Q-Bond, the primer and polymerase bind sequentially to the template, increasing primer annealing time.|
性能
The Rotor-Gene Probe PCR Kit is well suited for use in quantitative PCR analysis (see figures "Highly sensitive and reproducible detection", "High precision in real-time PCR"), and provides reliable detection over a wide range of templates (see table and figures "High precision in real-time PCR", "Reliable detection down to 1 cell", and "Wide dynamic range").
| 1,000,000,000 |
5.11 |
3.07 |
| 100,000,000 |
8.18 |
3.39 |
| 10,000,000 |
11.57 |
3.04 |
| 1,000,000 |
14.61 |
3.68 |
| 100,000 |
18.29 |
3.28 |
| 10,000 |
21.57 |
3.68 |
| 1,000 |
25.25 |
3.06 |
| 100 |
28.31 |
3.43 |
| 10 |
31.74 |
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| Efficiency |
0.99 |
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| Linearity |
1 |
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原理
The Rotor-Gene Probe PCR Kit enables reliable real-time two-step RT-PCR quantification on the Rotor-Gene Q without the need for optimization of reaction and cycling conditions. The kit is designed for use with hydrolysis probes (e.g., TaqMan® probes). Highly specific amplification is assured through a balanced combination of K+ and NH4+ ions, which promote specific primer annealing, enabling high PCR specificity and sensitivity (see figure "Specific primer annealing"). Fast cycling without compromising performance is achieved using Q-Bond, a novel PCR additive that enables cycler run times of as low as 45 minutes (see figure "Fast primer annealing").
| HotStarTaq Plus DNA Polymerase |
5 min activation at 95ºC |
Set up of qPCR reactions at room temperature |
| Rotor-Gene Probe PCR Buffer |
Balanced combination of NH4+ and K+ ions |
Specific primer annealing ensures reliable qPCR results |
| Unique Q-Bond additive |
Faster PCR run times enable faster results and more reactions per day |
操作流程
The ready-to-use Rotor-Gene Probe PCR Master Mix eliminates the need for optimization of reaction and cycling conditions. Just add template DNA, primers, and probe to the master mix and program the cycler. Instructions are provided in the detailed handbook supplied with the kit.
Hydrolysis probes (e.g., TaqMan® probes) can be used in combination with the Rotor-Gene Probe PCR Kit on the Rotor-Gene Q for fast and sensitive quantification — simply add the primer-probe mix and template to the master mix.
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.
应用
The Rotor-Gene Probe PCR Kit is for fast, real-time PCR and two-step RT-PCR of gDNA or cDNA targets using sequence-specific probes on the Rotor-Gene Q. It is also compatible with Rotor-Gene 3000 and the Rotor-Gene 6000 cyclers.
For ultrafast, one-step qRT-PCR gene expression analysis of RNA targets using sequence-specific probes on Rotor-Gene cyclers, use the Rotor-Gene Probe RT-PCR Kit.
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特点
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参数
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应用
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Real-time quantification of genomic DNA or cDNA targets
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描述
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For ultrafast quantitative real-time PCR and two-step RT-PCR using sequence-specific probes
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反应类型
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Real-time PCR and two-step RT-PCR
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real-time或终点法PCR
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Real-time
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样本/目标类型
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DNA, cDNA
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单一或多重
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Single
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SYBR Green I或序列特异性探针
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Sequence-specific probes
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热循环仪
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Rotor-Gene Q, Rotor-Gene 3000, Rotor-Gene 6000
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有/无ROX
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Without ROX dye
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For fast real-time PCR, two-step RT-PCR, and one-step RT-PCR using sequence-specific probes on Rotor-gene cyclers.
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图片
灵敏、可重复的检测。
使用2倍稀释的基因组DNA（30 ng到3.75 ng）和自行设计的TaqMan探针对IL1R2（白介素1受体Ⅱ）进行real-time PCR。反应使用[A] Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR仪或[B]供应商AII提供的试剂盒和循环仪（每个稀释样本重复五次反应）。Rotor-Gene体系能够检测到较低的CT值，有更好的重复性以及对不同浓度模板的较高的分辨率。
real-time PCR准确度高。
使用Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR分析仪对人基因组DNA进行real-time PCR分析。[A] 2倍稀释的DNA（从30 [5000拷贝]到0.06 ng[20拷贝]）使用自行设计的TaqMan探针对IL1R2（白介素1受体Ⅱ）进行分析；每个浓度模板重复五次。相邻浓度模板间的CT值平均差1.07个循环。[B] 5 ng DNA使用自行设计的TaqMan探针对IL1R2进行分析。100次重复实验的标准偏差为0.21。[C] 20 ng DNA使用自行设计的TaqMan探针对ACTB（β-actin）进行分析。100次重复实验的标准偏差为0.29。
对1个细胞也能可靠检测。
使用4倍稀释的人基因组DNA（相当于从16, 666个细胞到1个细胞）和自行设计的TaqMan探针对ACTB（β-actin）进行real-time PCR分析。反应使用Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR分析仪进行，重复三次实验。Rotor-Gene体系在整个浓度范围内检测可靠且重复性好。
宽泛的线性范围。
使用10倍稀释的质粒DNA（109 到10个拷贝）和自行设计的TaqMan探针对PPIA（亲环素A）进行real-time PCR分析。反应使用Rotor-Gene Probe PCR Kit和Rotor-Gene Q实时荧光定量PCR分析仪进行，重复三次实验。Rotor-Gene体系在整个浓度范围内检测可靠且CT值重复性好。
Specific primer annealing.
Cations in the Rotor-Gene Q PCR buffer increase specific primer annealing. K+ binds to phosphate groups on double-stranded DNA, stabilizing primer annealing. NH4+ destabilizes weak hydrogen bonds between mismatched bases.
Fast primer annealing.
[A] Q-Bond in Rotor-Gene Probe PCR Master Mix increases the affinity of DNA polymerase for short single-stranded DNA, reducing primer annealing time to a few seconds. In addition, the unique buffer composition supports the melting of DNA, reducing denaturation and extension times. [B] Without Q-Bond, the primer and polymerase bind sequentially to the template, increasing primer annealing time.
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