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。

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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
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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
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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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