For highly sensitive detection of viral RNA and/or DNA
- High sensitivity in single and multiplex assays
- Detection of viral RNA and/or DNA in the same reaction
- Clear detection of weak positive signals
- Fast universal 2-step protocols
- 5x master mix for higher sensitivity with more sample input
The QuantiTect Virus Kit is specially designed for highly sensitive detection of up to 4 viral nucleic acid targets by multiplex, real-time one-step RT-PCR using sequence-specific probes. Multiple viral RNA and/or DNA targets plus internal controls can be detected in a single tube without loss of sensitivity. The concentrated 5x master mix enables the addition of higher sample input and contains HotStarTaq Plus DNA Polymerase. The QuantiTect Virus RT Mix contains a unique formulation of Sensiscript Reverse Transcriptase that is optimized for highly sensitive detection of viral RNA. Two kit formats are available: the QuantiTect Virus Kit for cyclers that require ROX dye for fluorescence normalization and the QuantiTect Virus +ROX Vial Kit for other cyclers. For convenience, the master mix can be stored at 2–8°C.
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QuantiTect Virus Kit (50)
Trial kit for 50 x 50 µl reactions: QuantiTect Virus Master Mix (contains ROX dye), QuantiTect Virus RT Mix, RNase-Free Water, QuantiTect Nucleic Acid Dilution Buffer
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211011
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QuantiTect Virus Kit (200)
For 200 x 50 µl reactions: QuantiTect Virus Master Mix (contains ROX dye), QuantiTect Virus RT Mix, RNase-Free Water, QuantiTect Nucleic Acid Dilution Buffer
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211013
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QuantiTect Virus Kit (1000)
For 1000 x 50 µl reactions: QuantiTect Virus Master Mix (contains ROX dye), QuantiTect Virus RT Mix, RNase-Free Water, QuantiTect Nucleic Acid Dilution Buffer
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211015
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QuantiTect Virus +ROX Vial Kit (200)
For 200 x 50 µl reactions: QuantiTect Virus NR Master Mix (without ROX dye), ROX Dye Solution, QuantiTect Virus RT Mix, RNase-Free Water, QuantiTect Nucleic Acid Dilution Buffer
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211033
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QuantiTect Virus +ROX Vial Kit (1000)
For 1000 x 50 µl reactions: QuantiTect Virus NR Master Mix (without ROX dye), ROX Dye Solution, QuantiTect Virus RT Mix, RNase-Free Water, QuantiTect Nucleic Acid Dilution Buffer
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211035
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QuantiTect Virus Kits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。
准确测定CT值。|可靠的检测病毒RNA。|QIAGEN多重PCR试剂盒。|独特的PCR缓冲液。|更好的检测少量病毒RNA。|对RNA标准品进行可靠的稀释和储存。|
如图所示,使用QuantiTect Virus Kit扩增并分析梯度稀释的牛病毒性腹泻病毒(BVDV)RNA。即便是低模板量(10个拷贝/μl),陡峭的s形曲线亦可实现准确的CT值测定。[A] BVDV1 RNA的检测。[B] BVDV2 RNA的检测。NTC:无模板对照。|使用QuantiTect Virus Kit双重扩增5倍梯度稀释的病毒RNA及内部对照。[A] 病毒靶点的扩增曲线。[B] 内对照的扩增曲线。|QuantiTect Virus Kit提供了简单的多重定量real-time PCR步骤。与当前的方法相比,即便是进行复杂的分析(如目的基因的拷贝数明显少于对照基因的分析),该试剂盒亦无需优化引物、Mg2+和Taq DNA聚合酶的浓度。|[A] NH4+防止引物与模板的非特异性结合。[B] MP合成因子是创新的PCR添加剂,能够提高模板处局部的引物浓度,与K+和其他离子一起,MP合成因子能够特异性稳定结合的引物,使HotStarTaq Plus DNA Polymerase作用下的引物延伸更加高效。|使用QuantiTect Virus Kit或Suppliers AII和I的试剂盒双重扩增5倍梯度稀释的病毒RNA及内对照。QuantiTect Virus Kit的灵敏度大大高于其他检测试剂盒,可实现未知样本的可靠分析。ND:在50个PCR循环后未检测到。|使用QuantiTect Nucleic Acid Dilution Buffer或不含RNA酶的纯水按说明制备连续10倍梯度稀释的RNA标准品(体外转录的RNA)。直接使用这些稀释样本作为模板或将其置于-20°C保存2或4周后进行一步法RT-PCR。使用QuantiTect Nucleic Acid Dilution Buffer稀释标准品可获得更低的CT值和更佳的稳定性。|
性能
Amplification using QuantiTect Virus Kits provides steep sigmoidal curves over a range of dilutions, even for low template amounts with high CT values (see figure "Unambiguous determination of CT values over a wide dynamic range"). This enables accurate CT value determination for quantification of viral nucleic acids in real-time PCR.
Multiplex assays enable the detection of multiple viral RNA and/or DNA targets plus internal controls over a wide linear range without loss of sensitivity (see figures "Reliable detection of viral RNA over a wide linear range" and "Improved detection of low amounts of viral RNA").
QuantiTect Nucleic Acid Dilution Buffer, supplied with the kits, stabilizes RNA and DNA standards during dilution and reaction setup and prevents loss of nucleic acids on plastic surfaces, such as tubes or pipet tips. The buffer enables reliable dilution of standards used to quantify viral nucleic acids, giving a wide linear range, from low to high CT values and ensures longer storage of standards without degradation (see figure "Reliable dilution and storage of RNA standards").
原理
QuantiTect Virus Kits provide highly sensitive detection of viral nucleic acids in single or multiplex assays on the first attempt (see flowchart "QIAGEN multiplex kits"). The optimized master mix ensures that PCR products in a multiplex reaction are amplified with the same efficiency and sensitivity as PCR products in a corresponding single-amplification reaction.
Amplifying control and target genes in the same reaction, instead of separate reactions, increases the reliability of gene quantification by minimizing handling errors. The QuantiTect Virus Buffer contains a balanced combination of K+ and NH4+ ions as well as unique synthetic Factor MP stabilizes, which together promote stable and efficient annealing of primers and probes to the nucleic acid template, enabling high PCR efficiency (see figure "Unique PCR buffer"). In addition, the unique formulation of Sensiscript Reverse Transcriptase ensures highly sensitive reverse transcription of viral RNA, while HotStarTaq Plus DNA Polymerase provides a stringent hot start, preventing the formation of nonspecific products.
| 5x QuantiTect Virus Master Mix |
Concentrated master mix |
Highly concentrated and optimized for sensitive virus detection |
Larger volumes of template can be added to the assay for increased sensitivity |
| HotStarTaq Plus DNA Polymerase |
5 min activation at 95ºC |
Set up of qPCR reactions at room temperature |
| QuantiTect Virus Buffer |
Balanced combination of NH4+ and K+ ions |
Specific primer annealing ensures reliable PCR results |
| Synthetic Factor MP |
Reliable multiplexing analysis of up to 4 genes in the same tube |
| Additional kit components |
QuantiTect Virus RT Mix |
Contains a unique formulation of Sensiscript Reverse Transcriptase |
Optimized for highly sensitive detection of viral RNA |
| QuantiTect Nucleic Acid Dilution Buffer |
Proprietary buffer formulation for dilution and storage of nucleic acid standards. |
Stabilizes RNA and DNA standards during dilution and reaction setup and prevents loss of nucleic acids on plastic surfaces, such as tubes or pipet tips |
操作流程
QuantiTect Virus Kits provide highly sensitive real-time PCR analysis of viral nucleic acids (RNA and/or DNA) and internal controls using sequence-specific probes. Reactions can be carried out with or without a reverse-transcription step, enabling flexible design of multiplex assays to detect RNA targets, DNA targets, or both RNA and DNA targets. Follow the protocol in the handbook for fast and reliable results.
Kits are available with or without ROX passive reference dye in the master mix (see table).
| Supplied in master mix |
QuantiTect Virus Kit |
All cyclers from Applied Biosystems except Applied Biosystems 7500 |
| Supplied in separate tube |
QuantiTect Virus +ROX Vial Kit |
Applied Biosystems 7500 and cyclers from Bio-Rad, Cepheid, Eppendorf, QIAGEN, Roche, Agilent, and other suppliers |
For fast and highly sensitive end-point one-step RT-PCR applications, including virus detection, we recommend using the QIAGEN OneStep RT-PCR Kit.
应用
QuantiTect Virus Kits provide highly sensitive real-time single or multiplex PCR or one-step RT-PCR using sequence-specific probes for detection of viral DNA and/or RNA plus internal controls. The kits can be used on a wide range of real-time thermal cyclers, including instruments from Applied Biosystems, Bio-Rad, Cepheid, Eppendorf, QIAGEN, Roche (except for capillary cyclers), and Agilent.
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特点
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参数
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应用
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Virus detection
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反应类型
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Reverse Transcription and 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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RNA and/or DNA targets
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单一或多重
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Single or multiplex
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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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Most real-time cyclers (except capillary cyclers e.g. LightCycler® 1.x and 2.0)
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有/无ROX
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Available with ROX in master mix and ROX as a separate vial
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Now with even more applications!
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FAQ ID -2448
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FAQ ID -2451
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FAQ ID -2682
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浏览
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FAQ ID-751
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浏览
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For highly sensitive real-time singleplex or multiplex PCR or one-step RT-PCR using sequence-specific probes for detection of viral DNA and RNA and internal controls
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图片
准确测定CT值。
如图所示,使用QuantiTect Virus Kit扩增并分析梯度稀释的牛病毒性腹泻病毒(BVDV)RNA。即便是低模板量(10个拷贝/μl),陡峭的s形曲线亦可实现准确的CT值测定。[A] BVDV1 RNA的检测。[B] BVDV2 RNA的检测。NTC:无模板对照。
可靠的检测病毒RNA。
使用QuantiTect Virus Kit双重扩增5倍梯度稀释的病毒RNA及内部对照。[A] 病毒靶点的扩增曲线。[B] 内对照的扩增曲线。
QIAGEN多重PCR试剂盒。
QuantiTect Virus Kit提供了简单的多重定量real-time PCR步骤。与当前的方法相比,即便是进行复杂的分析(如目的基因的拷贝数明显少于对照基因的分析),该试剂盒亦无需优化引物、Mg2+和Taq DNA聚合酶的浓度。
独特的PCR缓冲液。
[A] NH4+防止引物与模板的非特异性结合。[B] MP合成因子是创新的PCR添加剂,能够提高模板处局部的引物浓度,与K+和其他离子一起,MP合成因子能够特异性稳定结合的引物,使HotStarTaq Plus DNA Polymerase作用下的引物延伸更加高效。
更好的检测少量病毒RNA。
使用QuantiTect Virus Kit或Suppliers AII和I的试剂盒双重扩增5倍梯度稀释的病毒RNA及内对照。QuantiTect Virus Kit的灵敏度大大高于其他检测试剂盒,可实现未知样本的可靠分析。ND:在50个PCR循环后未检测到。
对RNA标准品进行可靠的稀释和储存。
使用QuantiTect Nucleic Acid Dilution Buffer或不含RNA酶的纯水按说明制备连续10倍梯度稀释的RNA标准品(体外转录的RNA)。直接使用这些稀释样本作为模板或将其置于-20°C保存2或4周后进行一步法RT-PCR。使用QuantiTect Nucleic Acid Dilution Buffer稀释标准品可获得更低的CT值和更佳的稳定性。
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