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 Pfu Plus 5x PCR Master Mix

 

Products

Qty.

Cat. No.

Conc.

Price(W)

Protocol

MSDS

 Pfu Plus 5x PCR Master Mix

1 ml

EBT-1403

250 rxn/20ml

120,000

5x1 ml

1,250 rxn/20ml

480,000

 

Description

Pfu Plus 5x PCR Master Mix is a ready-to-use premix containing all the components essential for a PCR and agarose gel electrophoresis (DNA polymerase, dNTP, reaction buffer, glycerol, bromophenol blue, and stabilizer). PCR can be performed simply by adding primer pair and template.

As Master Mix is supplied as a 5x concentration format, users should adjust a final reaction to 1x (if final reaction volume is 20 ml, 4 ml of 5x Master Mix should be added). One unit of Pfu Plus DNA polymerase is contained in 4 ml of 5x Master Mix.

Pfu Plus DNA Polymerase is suitable for a reliable amplification of long (up to 12 kbp) and complex targets with a robust yield and high accuracy. Although the error rate is slightly lower than that of Pfu DNA polymerase, the amplification efficiency is about 3-5 times more. The amplified products by Pfu Plus DNA Polymerase can be used for a cloning of your genes with a decreased error rate, and highly useful for a site-specific mutagenesis. Pfu Plus DNA Polymerase, like any other polymerases showing proof-reading activity, generates a PCR product with blunt end.

Pfu Plus DNA Polymerase exhibits 3' to 5´ exonuclease (proof-reading) activity, but has no detectable 5' to 3´ exonuclease activity.

QC tests

Performance tests, genomic DNA contamination test, confirmation test for the absence of endo and exonucleases.

Storage Condition

Store at -20°C.

Pfu Plus 5x PCR Master Mix is stable for at least 2 years at recommended storage condition.

Standard Protocol

1.Prepare 20 ml PCR solution as follows:

  PCR grade distilled water :   - ml

  Pfu Plus 5x PCR Master Mix :   4 ml

  Primer (10 pmol/ml) :   0.5 ml each

  Template DNA :  0.1-10 ng

  Adjust final vol. to 20 ml with PCR grade distilled water

2. Set PCR cycling as follows :

 Initial denature at 95°C : 3 min

 

1-2 kbp  

3-5 kpb 

> 6 kbp

Denature  : 95°C 

20 sec

20 sec

30 sec

Anneal  : Tm-4°C

20 sec

20 sec

30 sec 

Extend  : 72°C 

30 sec

30 sec/kbp

30-60 sec/kbp

* Set 25-35 PCR cycles for effective amplification :

  20-25 cycles for plasmid DNA, 30-40 cycles for gDNA, 35-45 cycles for RT products

* You can also use two step cycle amplification (denaturation at 95°C and annealing/extension at 68°C)

3. You can analyze PCR products by direct loading into agarose gel because PCR Master Mix contains glycerol and bromophenol blue (blue color) essential for a gel loading.

 

 

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