PCR & DNA modifying enzymes

DF Taq DNA Polymerase Basic kit, 1000 U

DF Taq DNA polymerase is a thermostable DNA polymerase manufactured with the highest standards. It is certified as DNA free (DF) as well as RNase and DNAse free, that guarantee reproducible and efficient PCR reactions.

Reagents supplied: DF Taq DNA polymerase, 10x Taq DNA polymerase Buffer with 15mM MgCl2, 10x Taq DNA polymerase Buffer w/o MgCl2, 25mM MgCl2

Unit definition: One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmoles of dNTPs into acid insoluble material in 30 minutes at 72oC.

(SKUs: PD010S)

 

For detailed product information please refer to the IFU

Product Details

Taq DNA Polymerase is a thermostable enzyme that catalyzes 5’→3′ synthesis of DNA. The enzyme has no detectable 3’→5′ proofreading exonuclease activity but possesses low 5’→3′ exonuclease activity. DF Taq DNA Polymerase has high processivity up to 5 kb, while it can reliably amplify a substrate as low as 6-12 copies per reaction e.g. 16 S rRNA fragment is consistently detected by PCR containing 1.25 units of DF Taq DNA polymerase in reactions where 10 fg of E.coli genomic DNA standard is added.

PCR Guidelines:

PCR recommended reaction:

Components

25μl assay

Final Concentration/

Quantity (25μl assay)

50μl assay

Final Concentration/

Quantity (50μl assay)

10x Taq pol. Buffer with 15mM MgCl2

2.5μl

1x

5μl

1x

dNTP mix 10mM each

0.5μl

0.2 mM each

1μl

0.2 mM each

10μΜ forward primer

0.5μl

0.2-0.5 μM

2.5μl

0.2-0.5 μM

10μΜ reverse primer

0.5μl

0.2-0.5 μM

2.5μl

0.2-0.5 μM

Template DNA

Variable

10 fg-500 ng*

Variable

100 fg-500 ng*

DF Taq DNA pol. (5u/μl)

0.25μl

1.25 u/25 μl reaction

(0.05 u/μl)

0.25-0.5μl

1.25-2.5 u/50 μl reaction

(0.025-0.05 u/μl)

Sterile ultrapure water

Up to 25 μl

 

Up to 50 μl

 

* Substrate quantity is mostly dependent on the complexity and the purity of substrate and should be defined after testing for each primer set/substrate combination. For highly pure DNA substrates (OD260/OD280 ~ 2 and OD260/OD230~ 2,2) dissolved in water for molecular biology, 5-10 copies of specific target substrate are enough for end product gel visualization.

PCR recommended conditions:

Step

Temperature

Time

Initial denaturation

95oC

5 min

25-35 cycles

Denaturation

95oC

30sec

Annealing

45-68oC*

20sec

Extension

72oC

1min/kb

 

 

 

Final extension

72oC

5 min

Hold

4oC

Indefinitely

*Annealing temperature depends on primers’ Tm

Functional Quality Control:

DF Taq DNA Polymerase is tested for performance in the polymerase chain reaction (PCR) using 1.25 units of enzyme to amplify:

  1. a 400-bp region of coli ribosomal DNA using 100 fg of E.coli gDNA as substrate in a 25μl reaction. The resulting PCR product is visualized as a single band on agarose gel.
  2. a 5000-bp region of Lambda DNA using 20ng as substrate in a 50μl reaction. The resulting PCR product is visualized as a single band on agarose gel.

Other Quality Controls:

Certified as DNase-RNase free using fluorescently labeled RNase and DNase probes. DNA free contamination is tested extensively through PCR for the detection of E.coli 16S ribosomal DNA. More details concerning quality controls could be found in respective Certificate of analysis.

Shipping

Shipped on blue ice or dry ice

Storage conditions

Store at -20οC ± 5οC

Shelf life:

12 months upon production

title

Go to Top