Description

 

  Real-Time qPCR 2x Master Mix includes all kind of real-Time qPCR components such as hot start Taq DNA polymerase, dNTP and buffer. The only need to perform the reactions template DNA, Taqman probes and primers provided by users.

 

  Especially, this product is suitable to perform the Taqman real-time qPCR reaction, since it has Taq DNA polymerase having the 5’ to 3’ exonuclease activity.

 

  This product has capable of performing the accurate quantitative analysis with high sensitivity and broad dynamic range using variants templates. Because this product includes all the components except template, primers and probes, it is very easy to use and reduce significantly the fault and bias. 

 

Applications

 

  - Real-Time quantification of DNA and cDNA targets.

 

  - Gene expression profiling.

 

  - Microbial & viral pathogen detection.

 

  - Diverse applications using qPCR 

 

Qualifying Test

 

  Performance test, exo/endo nuclease contamination test, stability test. 

 

Storage Condition

 

  For long term storage, we recommend to store at -20℃ (stable up to 6 months), and store at 4℃ after thawing the solution instead of -20℃. 

 

Standard Experiment Protocol

1. Thaw the qPCR 2x Master Mix.

2. Prepare the 20㎕ of Real-Time qPCR reaction mix as follow.

 

2. Prepare the 20㎕ of Real-Time qPCR reaction mix as follow.

 

qPCR 2x Master Mix                                  10 ㎕ 

Primer up-stream (10 pmol/㎕)            0.5 - 1 ㎕

Primer down-stream (10 pmol/㎕)       0.5 - 1 ㎕

Taqman probe                                     0.5 - 1 ㎕

Template DNA                                       variable

 

Add distilled water to make 20 ㎕ final volume.

    

3. After mix and put the tubes or plates.

4. Seal the tubes or plates with cap or optical adhesive film.

5. Run the Real-Time qPCR machine using the program.

Uracil-DNA glycosylase	50 ℃	5-10 min	UDG reaction
Initial denaturation	95 ℃	10 min	Hot start  Taq activation
Denaturation  Annealing & Extension	95 ℃  60 ℃	5-30 sec  30-60 sec	30-45 cycles

 

a. The number of PCR cycle is dependent on the copy number of DNA target in the samples.

 

b. Optimal annealing temperature is decided by the melting temperature of primer pairs. Usually, use the  2-4℃ below than the melting points of primer pairs.

 

c. Instead of two-step PCR setting in the above standard protocol, the three-step PCR cycle (denaturation, annealing and extension) can be used.

 

d. The precise adjustment of experimental procedure should be performed according to the samples and purpose.  

 

 

Performance Data