Cat. No.: HY-K0523

SYBR Green qPCR Master Mix (No ROX)

Name: SYBR Green qPCR Master Mix (No ROX)
Brand: MedChemExpress
SKU: HY-K0523
Price: 32.00 USD
Availability: InStock
Rating: 5 (42 reviews)

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Based on 42 publication(s) in Google Scholar

MCE SYBR Green qPCR Master Mix (No 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 no 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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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 (No ROX) can be directly used for robust and low-template quantitative PCR with high sensitivity, specificity and reliability. No ROX Reference Dye is included for use with instruments that does not require a reference dye for rectify the error of fluorescence signals between different wells.

MCE qPCR Master Mix (No ROX) is applied for the Real-Time PCR instrument as follows:

Bio-Rad: CFX96, CFX384, iCycler IQ, MyiQ, iQ5, MiniOpticon, Opticon, Opticon 2 and 4;

Eppendorf: Mastercycler ep realplex, realplex2 s;

Qiagen: Corbett Rotor-Gene Q, Corbett Rotor-Gene 3000, Corbett Rotor-Gene 6000;

Roche: LightCycler 480, LightCycler 2.0; LightCycler 96;

Thermo Scientific: PikoReal Cycler; Cepheid: SmartCycler; Illumina: Eco qPCR.

1. Ready to use.

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

3. High sensitivity, specificity and reliability.

4. No 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 (No 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. This formulation contains no ROX Reference Dye and is intended for instruments that do not require ROX normalization.

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.