Rotor-Gene SYBR® Green RT-PCR Kit

在Rotor-Gene PCR仪上使用SYBR Green I通过一步法qRT-PCR超快速进行基因表达分析

  • 经优化,可在Rotor-Gene PCR仪上超快速获得可靠的结果
  • 灵敏检测低拷贝靶基因
  • 精确检测广泛模板量范围内的基因
  • 特制的即用型预混液,用于快速循环
  • 配合QuantiTect Primer Assays使用,可保证性能

Rotor-Gene SYBR Green RT-PCR Kit专用于Rotor-Gene Q实时荧光定量PCR分析仪和其他的Rotor-Gene PCR仪,使用SYBR Green I对RNA靶基因进行超快速、高特异性的一步法real-time RT-PCR。经优化的预混液与独特的Rotor-Gene PCR仪配合使用保证了qPCR良好的性能。预混液可贮存在2–8°C,方便使用。

Product Cat. no. List price:
Rotor-Gene SYBR Green RT-PCR Kit (400)
For 400 x 25 µl reactions: 3 x 1.7 ml 2x Rotor-Gene SYBR Green RT-PCR Master Mix, 100 µl Rotor-Gene RT Mix, 2 x 2 ml RNase-Free Water
204174
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The Rotor-Gene SYBR® Green RT-PCR Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Specific detection without the need for optimization.|Specific primer annealing.|Fast primer annealing.|
Tenfold dilutions of human leukocyte RNA (100 ng to 10 pg) were used as template in SYBR® Green-based real-time one-step RT-PCR. Duplicate reactions were run using the QuantiTect Primer Assay for BCL2 (B-cell CLL/lymphoma 2). [A] The Rotor-Gene Q and Rotor-Gene SYBR® Green RT-PCR Kit provided sensitive detection from 10 pg RNA and amplification of specific PCR product (melting curve shown in inset). [B] In contrast, an instrument and kit from Supplier R provided detection only after optimization of Mg2+ concentration. However, the limit of detection was 100 pg RNA and coamplification of nonspecific products was observed (melting curve shown in inset).|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 SYBR® Green RT-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.|
Performance
QuantiTect Primer Assays与Rotor-Gene SYBR Green RT-PCR Kit联合使用,高灵敏度定量特异性PCR产物无需经过优化(参见"Specific detection without the need for optimization")。
Principle

Rotor-Gene SYBR Green RT-PCR Kit可在Rotor-Gene Q实时荧光定量PCR分析仪上进行快速、可靠的real-time RT-PCR定量分析,无需优化反应及循环条件。该产品利用一步法real-time RT-PCR技术,RNA在反应中作为模板,在同一反应体系中实现逆转录和PCR反应。因为无需将逆转录反应后的反应体系转移到另外的管中进行PCR反应,整个real-time RT-PCR反应十分高效,适用于高通量分析。

混合液中的荧光染料SYBR Green I能用于分析多种不同的靶基因而无需合成靶标特异性标签探针。预混液中浓度平衡的K+和NH4+离子组合保证了高特异性扩增,使引物退火高度特异性,获得高PCR特异性和灵敏性(参见"Specific primer annealing")。使用创新的PCR辅助剂Q-Bond使循环时间降低至45分钟,达到快速分析且不降低性能(参见"Fast primer annealing")。

2x Rotor-Gene SYBR Green RT-PCR Kit的组分*
组分 特点 优势
HotStarTaq Plus DNA Polymerase 在95ºC下5分钟活化 室温下进行qPCR反应体系构建
Rotor-Gene SYBR Green RT-PCR Buffer 浓度平衡的K+和NH4+离子组合 引物特异性结合保证可靠的qPCR结果
独特的Q-Bond添加剂 快速的PCR运行时间可获得较快的结果和每天更多的反应次数
SYBR Green I dye 与双链DNA结合产生较强的荧光信号 高度灵敏的定量检测
Rotor-Gene RT Mix 对RNA有高度亲和性的逆转录酶混合液 即便具有复杂的二级结构,RNA在10分钟内即可完成逆转录
* 同时含有dNTP预混液(dATP, dCTP, dGTP, dTTP)。
Procedure

即用型预混液无需优化反应及循环条件。只需将RNA模板、引物和逆转录酶混合液加入到预混液中并设置循环程序。在试剂盒使用手册中有详细说明。

对于基于real-time一步法RT-PCR的基因表达分析,Rotor-Gene SYBR Green RT-PCR Kit、QuantiTect Primer Assays及Rotor-Gene Q实时荧光定量PCR分析仪提供了完成的即用型解决方案。QuantiTect Primer Assays为覆盖全基因组,经功效验证的预制引物对,用于检测来自人、小鼠、大鼠和其他物种的转录本。QuantiTect Primer Assays可以便利的在GeneGlobe网上订购。

Applications

Rotor-Gene SYBR Green RT-PCR Kit适用于在Rotor-Gene Q实时荧光定量PCR分析仪上对RNA靶基因进行快速real-time定量分析。该试剂盒与Rotor-Gene 3000和Rotor-Gene 6000兼容。

Feature
Specifications
Applications Real-time quantification of RNA targets
Description For ultrafast quantitative real-time one-step RT-PCR using SYBR Green I
Reaction type Real-time one-step RT-PCR
Real-time or endpoint Real-time
Sample/target type RNA
Single or multiplex Single
SYBR Green I or sequence-specific probes SYBR Green I
Thermal cycler Rotor-Gene Q, Rotor-Gene 3000, Rotor-Gene 6000
With or without ROX Without ROX dye

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Brochures & Guides
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Kit Handbooks
2
For fast real-time PCR, two-step RT-PCR, and one-step RT-PCR using SYBR Green I on Rotor-Gene cyclers
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For genomewide, ready-to-use real-time RT-PCR assays using SYBR Green detection
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Safety Data Sheets
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Images
Specific and sensitive detection using SYBR Green
Specific detection without the need for optimization.
Tenfold dilutions of human leukocyte RNA (100 ng to 10 pg) were used as template in SYBR® Green-based real-time one-step RT-PCR. Duplicate reactions were run using the QuantiTect Primer Assay for BCL2 (B-cell CLL/lymphoma 2). [A] The Rotor-Gene Q and Rotor-Gene SYBR® Green RT-PCR Kit provided sensitive detection from 10 pg RNA and amplification of specific PCR product (melting curve shown in inset). [B] In contrast, an instrument and kit from Supplier R provided detection only after optimization of Mg2+ concentration. However, the limit of detection was 100 pg RNA and coamplification of nonspecific products was observed (melting curve shown in inset).
Specific primer annealing using a balanced combination of K+ and NH4+ ions
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 with Q-Bond
Fast primer annealing.
[A] Q-Bond in Rotor-Gene SYBR® Green RT-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.