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

STS-Pw250

STS-Pw1000

add nucleotides box

cat. no.

STSn-P250

STSn-P1000

amount

250 units

1000 units

250 units

1000 units

amount

250 units

1000 units

note

10X Standard Buffer

10X Standard Buffer

10X Standard Buffer w/o MgCl2

10X Standard Buffer

10X Standard Buffer w/o MgCl2

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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