基本描述

别名

FastSYBR 混合物

规格或纯度

5 ml

英文名称

FastSYBR Mixture

储存温度

-20°C储存

运输条件

超低温冰袋运输

产品介绍

FastSYBR Mixture是专用于染料法(SYBR GreenⅠ)实时荧光定量PCR的预混体系，浓度为2×,包含Fast Taq DNA Polymerase、PCR Buffer、dNTPs、SYBR Green I 荧光染料和Mg2+，操作简单方便。主要用于基因组DNA靶序列和RNA反转录后的 cDNA靶序列检测。本品所含荧光染料SYBR Green I可以与所有的双链DNA相结合，使该产品可用于不同靶序列的检测而不需合成特异性标记探针。本品含有的Fast Taq DNA Polymerase能有效减少在常温条件下由引物和模板非特异性结合或引物二聚体而产生的非特异性扩增，酶的激活仅需在95℃孵育20s。整个PCR反应过程比普通反应可节省约40分钟，大大缩短了PCR的反应时间。独特的PCR缓冲体系与热启动酶的组合，有效抑制了非特异产物的产生，并显著提高PCR的扩增效率。该产品应用范围广，适用于普通和快速定量PCR程序。　　ROX染料用于校正定量PCR仪孔与孔之间产生的荧光信号误差，一般用于ABI、 Stratagene等公司的Real Time PCR 扩增仪。不同仪器的激发光学系统有所不同，因此ROX染料的浓度必须与相应的荧光定量PCR仪相匹配。

不需要ROX校正的仪器： Roche LightCycler 480，Roche LightCyler 96，Bio-rad iCyler iQ，iQ5，CFX96等。

需要Low ROX校正的仪器： ABI Prism7500/7500 Fast，QuantStudio® 3 System，QuantStudio® 5 System， QuantStudio® 6 Flex System，QuantStudio® 7 Flex System，ViiA 7 system， Stratagene Mx3000/Mx3005P，Corbett Rotor Gene 3000等。

需要High ROX校正的仪器： ABI Prism7000/7300/7700/7900，Eppendorf，ABI Step One/Step One Plus等。

F665884	Component	5ml	Storage
F665884A	2×FastSYBR Mixture	5×1ml	-20℃. Avoid freeze/thaw cycle.
F665884B	ddH₂O	5×1ml	-20℃. Avoid freeze/thaw cycle.

注意事项

1．使用前请上下颠倒轻轻混匀，尽量避免起泡，并经短暂离心后使用。

2．本产品中含有荧光染料SYBR Green I，保存本产品或配制PCR反应液时应避免强光照射。

3．避免反复冻融本品，反复冻融可能使产品性能下降。本产品长期保存可置于-20℃，避光。如果在短期内需要频繁使用，可在2-8℃保存。

4．本品不能用于探针法荧光定量PCR。

使用方法

以下举例为常规PCR反应体系和反应条件，实际操作中应根据模板、引物结构和目的片段大小不同进行相应的改进和优化。

1. PCR反应体系

试剂	50 μL反应体系	终浓度
2×FastSYBR Mixture	25 μL	1 ×
Forward Primer，10 µM	1 μL	0.2 μM ¹⁾
Reverse Primer，10 µM	1 μL	0.2 μM ¹⁾
Template DNA	2 μl ²⁾	/
50×Low ROX or High ROX （optional）³⁾	1 μL	1 ×
ddH2O	up to 50 μl	/

注意：

1）通常引物浓度以0.2 μM可以得到较好结果，可以在0.1-1.0 μM作为设定范围的参考。扩增效率不高的情况下，可提高引物的浓度；发生非特异性反应时，可降低引物浓度，由此优化反应体系。

2）通常DNA模板的量以10-100 ng基因组DNA或1-10 ng cDNA为参照，因不同物种的模板中含有的目的基因拷贝数不同，可对模板进行梯度稀释，以确定最佳的模板使用量。

3）不同仪器的激发光学系统有所不同，根据使用荧光定量的仪器选择加入50×Low ROX or 50×High ROX.

2. PCR反应条件

步骤	温度	时间	/
预变性	95℃	20 s ¹⁾	/
变性	95℃	3 s	35-40 个循环
退火/延伸²⁾	60℃	30 s	35-40 个循环
融解曲线分析³⁾	/	/	/
/	95℃	15 s	/
/	60℃	1 min	/
/	95℃	15 s	/
/	60℃	15 s	/

注意：

1）本产品所采用的酶须在预变性95℃、20s条件下实现酶的活化。在此条件下，大多数模板可良好的进行解链。对GC含量高、二级结构复杂的模板，可将预变性时间延长至1分钟，以使起始模板充分解链。若高温处理时间过长，会对酶的活性造成影响。可根据模板情况确定最佳的预变性时间。

2）建议采用两步法PCR反应程序，退火温度请以60-64℃作为设定范围的参考，发生非特异性反应时，可提高退火温度。若因使用Tm值较低的引物等原因，得不到良好的实验结果时，可尝试进行三步法PCR扩增，退火温度请以 56℃-64℃的范围作为设定参考。

3）融解曲线分析请以所使用的荧光定量PCR仪推荐的程序进行设定，本程序是以ABI 7500荧光定量PCR仪为参照设定。

FastSYBR Mixture is a premixed system specifically designed for real-time fluorescence quantitative PCR using the dye method (SYBR Green I), with a concentration of 2 x, containing Fast Taq DNA Polymerase, PCR Buffer, dNTPs, SYBR Green I fluorescent dye, and Mg2+. The operation is simple and convenient. Mainly used for detecting genomic DNA target sequences and cDNA target sequences after RNA reverse transcription. The fluorescent dye SYBR Green I contained in this product can bind to all double stranded DNA, making it suitable for detecting different target sequences without the need for synthesizing specific labeled probes. The Fast Taq DNA Polymerase contained in this product can effectively reduce non-specific amplification caused by non-specific binding of primers and templates or primer dimers at room temperature. Enzyme activation only requires incubation at 95 ℃ for 20 seconds. The entire PCR reaction process can save about 40 minutes compared to ordinary reactions, greatly reducing the reaction time of PCR. The unique combination of PCR buffer system and hot start enzyme effectively inhibits the production of non-specific products and significantly improves the amplification efficiency of PCR. This product has a wide range of applications and is suitable for both regular and rapid quantitative PCR programs. ROX dye is used to correct the fluorescence signal error between wells in quantitative PCR instruments, and is generally used in Real Time PCR amplification instruments from companies such as ABI and Stratagene. The excitation optical systems of different instruments vary, so the concentration of ROX dye must be matched with the corresponding fluorescence quantitative PCR instrument.
Instruments that do not require ROX calibration: Roche LightCycle 480, Roche LightCylinder 96, Bio rad iCylinder iQ, iQ5, CFX96, etc.
Instruments that require Low ROX calibration: ABI Prism7500/7500 Fast, QuantStudio ® 3 System, QuantStudio ® 5 System, QuantStudio ® 6 Flex System, QuantStudio ® 7 Flex System, ViiA 7 System, Stratagene Mx3000/Mx3005P, Corbett Rotor Gene 3000, etc.
Instruments that require High ROX calibration include ABI Prism7000/7300/7700/7900, Eppendorf, ABI Step One/Step One Plus, etc.

Component	F665884 5 ml
2×FastSYBR Mixture	5×1 ml
50×Low ROX	/
50×High ROX	/
ddH2O	5×1 ml

Matters needing attention
1. Before use, please gently mix upside down to avoid foaming, and use after briefly centrifugation.
2. This product contains the fluorescent dye SYBR Green I. When storing this product or preparing PCR reaction solution, strong light exposure should be avoided.
3. Avoid repeated freezing and thawing of this product, as repeated freezing and thawing may cause a decrease in product performance. This product can be stored for a long time at -20 ℃, away from light. If frequent use is required in the short term, it can be stored at 2-8 ℃.
4. This product cannot be used for probe based fluorescence quantitative PCR.
Usage
The following are examples of conventional PCR reaction systems and reaction conditions. In practical operation, corresponding improvements and optimizations should be made based on different templates, primer structures, and target fragment sizes.
1. PCR reaction system

Reagent	50 μL Reaction system	Final concentration
2×FastSYBR Mixture	25 μL	1 ×
Forward Primer，10 µM	1 μL	0.2 μM ¹⁾
Reverse Primer，10 µM	1 μL	0.2 μM ¹⁾
Template DNA	2 μl ²⁾	/
50×Low ROX or High ROX （optional）³⁾	1 μL	1 ×
ddH2O	up to 50 μl	/

Attention:
1) Typically, the primer concentration is 0.2 μ M can achieve good results, ranging from 0.1 to 1.0 μ M serves as a reference for setting the range. In the case of low amplification efficiency, the concentration of primers can be increased; When non-specific reactions occur, the primer concentration can be reduced to optimize the reaction system.
2) The amount of DNA template is usually based on 10-100 ng genomic DNA or 1-10 ng cDNA as a reference. Due to the different copy numbers of target genes contained in templates of different species, gradient dilution can be applied to the template to determine the optimal template usage.
3) The excitation optical systems of different instruments vary, and 50 x Low ROX or 50 x High ROX can be added according to the instrument used for fluorescence quantification
2. PCR reaction conditions

Step	Temperature	Time	/
Pre denaturation	95℃	20 s ¹⁾	/
Denaturation	95℃	3 s	35-40 cycles
Annealing/Extension ² ⁾	60℃	30 s	35-40 cycles
Analysis of Fusion Curve ³ ⁾	/	/	/
/	95℃	15 s	/
/	60℃	1 min	/
/	95℃	15 s	/
/	60℃	15 s	/

Attention:
1) The enzyme used in this product must be activated under pre denaturation conditions of 95 ℃ and 20 seconds. Under these conditions, most templates can perform well in de chaining. For templates with high GC content and complex secondary structures, the pre denaturation time can be extended to 1 minute to fully unwind the starting template. If the high-temperature treatment time is too long, it will affect the enzyme activity. The optimal pre denaturation time can be determined based on the template situation.
2) It is recommended to use a two-step PCR reaction program, and the annealing temperature should be set at 60-64 ℃ as a reference range. When non-specific reactions occur, the annealing temperature can be increased. If good experimental results cannot be obtained due to the use of primers with lower Tm values, a three-step PCR amplification method can be attempted. The annealing temperature should be set within the range of 56 ℃ -64 ℃ as a reference.
3) Please set the fusion curve analysis according to the recommended program of the fluorescence quantitative PCR instrument used. This program is based on the ABI 7500 fluorescence quantitative PCR instrument as a reference setting.

安全和危险性（GHS）

SDS英文版

技术规格说明书

技术规格说明书

f665884_fastsybr mixture.pdf

质检证书（CoA,COO,BSE/TSE 和分析图谱)

C of A & Other Certificates(BSE/TSE, COO):

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FastSYBR Mixture

库存信息

基本描述

安全和危险性（GHS）

技术规格说明书

质检证书（CoA,COO,BSE/TSE 和分析图谱)

溶液计算器

摩尔浓度计算器

稀释计算器

复溶计算器