GeneSpin Srl
Via Friuli, 51
20135 Milano
P.IVA C.F. 04520270960
Laboratory
and sales office
c/o Fondazione Filarete
Viale Ortles, 22/4
20139 Milano
tel. +39 02 56660199
fax. +39 02 537250
Pfu DNA Pol is a superior thermostable DNA Pol isolated from an E.coli strain carrying the “pol” gene from Pyrococcus furiosus with 5’ →3’ polymerase activity and 3’ → 5’ exonuclease proofreading activity, i.e. it catalyzes DNA synthesis with very low error rate (about twelve times more accurate than Taq Pol). However, Pfu DNA Pol is slower than Taq Pol and typically requires about 1-2 min per cycle to amplify 1Kb of DNA.Using Pfu DNA Pol in PCR reactions also results in blunt-ended amplification products. Pfu DNA Pol is hence superior for techniques that require high fidelity DNA synthesis, but can also be used in conjunction with Taq Pol to obtain the fidelity of Pfu with the speed of Taq Pol activity.
cat. no.
STS-P250
STS-P1000
amount
250 units
1000 units
note
10X Standard Buffer
10X Standard Buffer
STS-Pw250
STS-Pw1000
250 units
1000 units
10X Standard Buffer w/o MgCl2
10X Standard Buffer
10X Standard Buffer w/o MgCl2
add nucleotides box
cat. no.
STSn-P250
STSn-P1000
amount
250 units
1000 units
note
10X Standard Buffer + dNTPs
10X Standard Buffer + dNTPs
STSn-Pw250
STSn-Pw1000
250 units
1000 units
10X Standard Buffer w/o MgCl2 + dNTPs
10X Standard Buffer
10X Standard Buffer w/o MgCl2 + dNTPs
FOR RESEARCH USE ONLY
UNIT DEFINITION
One unit is defined as the amount of enzyme required to incorporate 10 nanomoles of dNTPs into acid-insolubile material in 30 min at 74°C.
SHIPPING
Shipped in green ice.
STORAGE
Store at -20C°
SHELF LIFE
12 months
FORM
Liquid
CONCENTRATION
2U/ul
component
STS-P250
STS-Pw250
STSn-P250
STSn-Pw250
Pfu Polymerase
250 units /125ul
250 units /125ul
250 units / 125ul
250 units /125ul
Standard Buffer
1ml 10X Buffer with MgCl2
1ml 10X Buffer w/o MgCl2
1ml 10X Buffer with MgCl2
1ml 10X Buffer w/o MgCl2
MgCl2
-
500ul / 50mM
-
500ul / 50mM
dNTPs
-
-
100ul / 10mM each
100ul / 10mM each
component
STS-P1000
STS-Pw1000
STSn-P1000
STSn-Pw1000
Pfu Polymerase
1000 units /500ul
1000 units /500ul
1000 units /500ul
1000 units /500ul
Standard Buffer
2ml 10X Buffer with MgCl2
2ml 10X Buffer w/o MgCl2
2ml 10X Buffer with MgCl2
2ml 10X Buffer w/o MgCl2
MgCl2
-
500ul / 50mM
-
500ul / 50mM
dNTPs
-
-
400ul / 10mM each
400ul / 10mM each
component
Standard Buffer
dNTPs
Pfu Polymerase
primers
DNA Template
Assay Set-Up:
Before starting, vortex all components thoroughly to ensure homogeneity.
Prepare a premix for the number of assays you need according to the following protocol:
stock conc.
10X
final conc.
1X
30ul reaction
10mM each
200uM
2U/ul
0.5U/ul
1ug/ul each
50ng/ul each
-
-
3.0ul
0.6ul
1.0ul
2.0ul each
10-20ng
1)The annealing temperature depends on the melting temperature of the primers used.
2)The elongation time depends on the length of the fragments to be amplified. A time of 1 min/kb is recommended.
Cycling conditions:
Spin down the tubes/plate briefly to remove bubbles and place them into the cycler.
denaturation
denaturation (1)
annealing (2)
extension
95°C
95°C
50-68°C
72°C
5min
30 sec
30 sec
30 sec
4 min
MG Water
-
-
up to 30ul
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