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Cat. No.: HY-K0521

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SYBR Green qPCR Master Mix (High ROX)

Name: SYBR Green qPCR Master Mix (High ROX)
Brand: MedChemExpress
SKU: HY-K0521
Price: 32.00 USD
Availability: InStock
Rating: 5 (5 reviews)

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MCE SYBR Green qPCR Master Mix (High ROX) is provided as a simple-to-use, stabilized 2× formulation that includes all components for qPCR except sample DNA, primers and water, in which High ROX Reference Dye is included. The 100 rxns is defined as the base specification. All larger sizes correspond to incremental volumes of this base.

Storage :
-20°C, 2 years. Protecting from light, and avoid repetitive freeze-thaw cycles.

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Fluorescent Dye

Strong Fluorescence Signal / Multi-Disciplinary Applications / High Fluorescence Specificity

Description & Advantages

The Super Hot-Start SYBR Green qPCR Master Mix is a ready-to-use qPCR cocktail. It contains a novel Taq DNA Polymerase, unique hot start reagents, optimized buffer, SYBR Green I, dNTPs, PCR Enhancer and PCR stabilizer. qPCR Master Mix is provided at 2× concentration and can be used at 1× concentration by adding template, primer and ddH₂O.

MCE qPCR Master Mix (High ROX) can be directly used for robust and low-template quantitative PCR with high sensitivity, specificity and reliability. High ROX Reference Dye is included for use with instruments that require a high level of reference dye for rectify the error of fluorescence signals between different wells.

MCE qPCR Master Mix (High ROX) is applied for the Real-Time PCR instrument as follows:

ABI GeneAmp 5700; ABI PRISM 7000, 7700; ABI 7300, 7900HT (Fast); ABI StepOne (Plus)

1. Ready to use.

2. Optimized buffer enhances specificity and reduces primer-dimer formation.

3. High sensitivity, specificity and reliability.

4. High ROX Reference Dye is included.

Protocol

1. Preparation of the reaction mixture: prepare the reaction mixture on ice. Three technical replicates are recommended for each sample.

Component	50 μL reaction	20 μL reaction	Final concentration
SYBR Green qPCR Master Mix (High ROX)	25 μL	10 μL	1×
Forward Primer (10 μM)	1 μL	0.4 μL	0.2 μM
Reverse Primer (10 μM)	1 μL	0.4 μL	0.2 μM
DNA/cDNA	4 μL	2 μL	/
ddH₂O	19 μL	7.2 μL	/
Total volume	50 μL	20 μL	/

Note: a. A final primer concentration of 200 nM is commonly used and may be optimized within 0.1-1.0 μM according to amplification efficiency and specificity;
b. Recommended template amount: 10-100 ng genomic DNA or 1-10 ng cDNA. Adjust the template amount according to target abundance to keep Ct values preferably within 15-35;
c. It is recommended to prepare a Master Mix without template first, dispense it, and then add template to reduce pipetting error.
d. High ROX Reference Dye is already included in the Master Mix; do not add extra ROX.

2. qPCR cycling program:

Step	Temperature	Time	Cycles
Initial denaturation	95°C	30 sec	1
Denaturation	95°C	10 sec	40
Annealing/Extension	60°C	30 sec	40
Melting curve	Use the instrument default program	-	-

3. Perform melting-curve analysis after amplification to confirm product specificity. Ramp rates and fluorescence acquisition settings may be adjusted according to the real-time PCR instrument used.