Super Pfx DNA Polymerase

Item Number
S665588
Grouped product items
SKUSizeAvailabilityPrice Qty
S665588-100U
100U
Available within 8-12 weeks(?)
Production requires sourcing of materials. We appreciate your patience and understanding.
$149.90
S665588-500U
500U
Available within 8-12 weeks(?)
Production requires sourcing of materials. We appreciate your patience and understanding.
$599.90
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Basic Description

Specifications & PurityEnzymoPure™
Storage TempStore at -20°C,Avoid repeated freezing and thawing
Shipped InIce chest + Ice pads
GradeEnzymoPure™
Product Description

Super Pfx DNA Polymerase is a fast and highly efficient high fidelity DNA polymerase with 5 ′ -3 ′ DNA polymerase activity and 3 ′ -5 ′ exonuclease activity. This enzyme is modified from other high fidelity enzymes, with strong amplification ability, fast amplification speed, and high fidelity, overcoming the shortcomings of poor amplification ability, low yield, and slow amplification speed of ordinary Pfu enzymes, greatly shortening the reaction time. This product can be used for long fragment amplification and other complex template amplification. The 3 'end of the PCR product obtained from amplification does not contain an "A" base and can be directly cloned into a flat end vector. If T/A cloning is required, "A" needs to be added to the end of the PCR product before cloning. This product is suitable for gene cloning, gene directed mutagenesis, SNP amplification experiments, etc.


S665588Component100 U500 UStorage
S665588ASuper Pfx DNA Polymerase, 2 U/μL50 μL250 μL-20℃. Avoid freeze/thaw cycle.
S665588B2×Super Pfx Buffer2×1.25 mL7×1.8 mL-20℃. Avoid freeze/thaw cycle.
S665588CdNTP Mix, 10mM each150 μL750 μL-20℃. Avoid freeze/thaw cycle.


Activity definition

The amount of enzyme required to add 10 nmoL deoxyribonucleotides to acidic insoluble substances within 30 minutes at 74 ℃ is defined as 1 active unit (U).

Quality control

After multiple column purifications, SDS-PAGE detected a purity of over 98%; No exogenous nuclease activity detected; Store at room temperature for one month without significant changes in activity.

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
All operations should be carried out on ice. After each group is decomposed and frozen, please mix thoroughly. After use, please put it back at -20 ℃ for storage in a timely manner.

Reagent

50 μLReaction system

final concentration

2×Super Pfx Buffer

25 μL

dNTP Mix,10 mM each

1.5-2.5 μL

300-500 μM each

Forward Primer,10 μM

2 μL

0.4 μM

Reverse Primer,10 μM

2 μL

0.4 μM

TemplateDNA appropriate amount

appropriate amount

<500 ng/50 μL

Super Pfx DNA  Polymerase

0.5-0.75 μL

1-1.5 U/50 μL

ddH2 O

up to 50 μL


2.  PCR reaction conditions

step

temperature

time

 

Pre denaturation

98°C

30 s-3 min

 

Degeneration regression

98°C

10-30 s

25-35 cycles

Fire extension

Determine based on primer Tm

15-30 s

25-35 cycles

Final extension

72°C

3-5 kb/min

25-35 cycles

extend

72°C

5 min


Attention:
1) Priority should be given to using the three-step method for amplification. If the three-step method cannot amplify the target product or the Tm value of the primer is greater than 68 ° C, please try the two-step method.
2) Denaturation: Pre denaturation of simple templates at 98 ℃ for 30 seconds to 1 minute. For complex templates, the pre denaturation time can be extended to 3 minutes.
3) Annealing: In general experiments, the annealing temperature is 3-5 ℃ lower than the Tm value of the primer. If the ideal amplification efficiency cannot be achieved, the annealing temperature should be gradually changed for optimization; When non-specific reactions occur, increase the annealing temperature appropriately.
4) Extension: The extension time should be set according to the length of the amplified fragment and the complexity of the template. The amplification efficiency of this product is 3-5 kb/min, and it is recommended to 2-4 kb/min for long fragments and high complexity templates.
5) Cycle count: The number of cycles can be set based on the downstream application of the amplification product. If there are too few cycles, insufficient amplification, or too many cycles, the probability of mismatch will increase. Therefore, while ensuring product yield, the number of cycles should be minimized as much as possible.

Certificates

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

Solution Calculators