|
Pfu
DNA Polymerase
Catalog
#
|
Pack
size
|
Price(€)
|
ZP00202
|
500U
|
14.40
|
ZP00203
|
1000U
|
27.20
|
Description:
Pfu DNA Polymerase is a thermostable
enzyme of approximately 90kDa isolated from Pyrococcus
furiosus. The enzyme replicates DNA at 75oC,
catalyzing the polymerization of nucleotides into duplex
DNA in the 5´-3´ direction in the presence
of magnesium. Pfu DNA Polymerase also possesses 3´-5´
exonuclease (proofreading) activity. Base misinsertions
that may occur during polymerization are rapidly excised
by the proofreading activity of the polymerase.
Consequently, Pfu DNA Polymerase is recommended for use
in PCR and primer extension reactions that require
high-fidelity synthesis. Pfu DNA Polymerase-generated
PCR fragments are blunt-ended.Elongation rate is 2Kbp/min.
Concentration:5U/ul
Source:
Pyrococcus furiosus strain Vc1 DSM3638.
5X Reaction Buffer with MgSO4:
100mM
Tris-HCl (pH 8.8 at 25oC),
50mM
KCl,
50mM
(NH4)2SO4,
10mM
MgSO4, 0.5% Triton X-100 and 0.5mg/ml
nuclease-free BSA.
Features:
High Fidelity: Pfu DNA Polymerase
exhibits the lowest error rate of any thermostable DNA
polymerase.
Applications:
Pfu DNA Polymerase is recommended for
use in PCR, primer extension reactions and other
applications that demand high fidelity.
Unit
Definition:
One unit is defined as the amount of
enzyme required to catalyze the incorporation of 10nmol
of dNTPs into acid-insoluble material in 30 minutes at
75oC.
Reaction
Mixture Set Up
1.
Gently vortex and briefly centrifuge all solutions after
thawing.
2.
Keep solutions on ice.
3.
Add to a thin-walled PCR tube, on ice:
Reagent
|
Quantity,
for 50 μl of reaction mixture
|
Final
concentration
|
5X Pfu buffer
|
10ul
|
1X
|
10mM
dNTP
|
1ul
|
0.2mM
|
PrimerI(25pmol/ul)
|
1ul
|
0.5pmol/ul
|
PrimerII(25pmol/ul)
|
1ul
|
0.5pmol/ul
|
Pfu(5U/ul)
|
0.4ul
|
2U/50ul
|
Template
DNA
|
variable
|
50
pg -1 μg
|
ddH2O
|
Up
to 50ul
|
|
4.
Gently vortex the sample and briefly centrifuge to
collect all drops from walls of tube.
5. If
using a thermal cycler without a heated lid, overlay the
sample with a half volume of mineral
oil
or add an appropriate amount of wax.
6.
Place samples in a thermal cycler preheated to
96°C
and start PCR.
Recommended
thermal cycling conditions:
|
Step
|
Temperature,°C
|
Time ,min
|
Number
of cycles
|
Initial
denaturation
|
96
|
4
|
1
|
Denaturation
|
95
|
0.5
|
|
Annealing
|
50-68
|
0.5-2
|
25-35
|
Extension
|
72
|
0.5-4
|
|
Final Extension
|
72
|
5
|
1
|
Quality
Control Tests:
PCR (activity), SDS-PAGE (purity), endonuclease/nickase.
Storage:
Pfu DNA Polymerase in
50mM
Tris-HCl (pH 8.2 at 25oC),
0.1mM
EDTA,
1mM
DTT, 50% glycerol and 0.05% CHAPS should be stored at
-20oC.Use
dry ice for shipping.
 Pfu
DNA Polymerase manual
If you want to get bulk order
price,please contact us.E-mail:master@shinegene.org.cn or shinegene@vip.163.com
Related Link
Peptide
Synthesis
Chromas
Make
Antibody
Lab
Consumables
Gene
Synthesis
dNTP
|
