DNA / PCR

Klenow-fragment of Taq DNA polymerase (KTQ)

(currently out of stock until further notice, please request)

 

is the perfect choice for standard PCRs and short amplicons. KTQ DNA polymerase supersedes the thermostability of Taq DNA polymerase, the relative mutation rate is twofold lower, but is less processive than Taq DNA polymerase and has no exonuclease or endonuclease activity.
KTQ DNA polymerase is supplied together with the 10x KTQ reaction buffer which has been specifically designed for optimal PCR performance and polymerase activity. KTQ DNA polymerase can also be used for realtime cycling, when adding a suitable realtime dye.

 

Article Product Size Price
#4001 S KTQ DNA polymerase 400 U, 5U/µl, 80 µl  € 45,-
#4001 M KTQ DNA polymerase 1000 U, 5U/µl, 200 µl  € 105,-
#4001 L KTQ DNA polymerase 4000 U, 5U/µl, 2x 400 µl  € 400,-

Bulk offer: Should you require larger amounts (> 10.000 U), significant discounts are available. Please contact us to receive an individual quotation.

 

download-pdf Product manual (PDF) Order

 

 

Example Results

Example Results

KTQ: Extreme thermostability2014_05_13

Comparison of  thermostability of KTQ and Taq DNA polymerase.

A fragment (92 bp) of the human actin was amplified from 10 ng of a human genomic DNA extract either with KTQ or Taq DNA polymerase with incubation at 95 – 97° C for one hour. PCR products were subsequently analysed on a 2.5% agarose gel.

 

Reaction Setup

Reaction Setup

Component Volume Final concentration
Primer forward (10 µM)* 1 µl 0.2 µM (0.05-1 µM)
Primer reserve(10 µM)* 1 µl 0.2 µM (0.05-1 µM)
dNTPs (2 mM) 5 µl 200 µM
10 x KTQ buffer 5 µl 1x
KTQ DNA polymerase 5 U/µl 0.25 µl 1.25 U / reaction
Template/Sample extract** x µl  
Nuclease-free water up to 50µl total reaction volume  

Keep all components on ice.
Spin down and mix all solutions carefully before use.
* Primers should ideally habe a GC-content of 40-60%
**Suggested template concentration should be about 1 ng - 1 µl (gnomic DNA) or 1 ng - 1 pg (plasmid/viral DNA).

Applications

Applications

  • Standard PCR
  • Realtime PCR
  • Primer extension reaction
  • TA cloning
  • 3`A-tailing of blunt ends
  • Screening / Hight-throughput PCRs

Typical PCR protocol

Typical PCR protocol

Step Temperature Time
Initial denaturation 95°C 2 min
Denaturation 95°C 15 sec
Annealing* 54-72°C 30 sec     (25-40 cycles)
Extension 72 °C 2 min / 1000 bp
Hold <10°C hold

*Typically, the annealing temperature is about 3-5°C below the calculated melting temperature of the primers used. 

 

Quality Control

PCR activity: KTQ DNA polymerase was tested for successful PCR performance. A 92 bp fragment (beta-actin gene) was amplified from human genomic DNA and analyzed by agarose gel electrophoresis.

DNA polymerase activity: KTQ DNA polymerase activity has been monitored and adjusted to a specific DNA polymerase activity using an artificial DNA template and a DNA primer.
Enzyme-concentration has been determined by protein-specific staining. Please inquire more information at info@mypols.de for the lot-specific concentration.
No contamination has been detected in standard test reactions.

 

Storage

This product is shipped on cool packs. Please store the product upon arrival at -20°C.

 

Composition / Content of the Product

KTQ DNA polymerase is supplied as a 5 U/µl solution containing glycerol. It comes together with an optimized 10x reaction buffer.

 

KTQ-teaser

 

img-faqsFAQ`s

Where can I read more about the KTQ DNA polymerase?

Have a look under following references for example:

The fidelity of Taq polymerase catalyzing PCR is improved by an N-terminal deletion.
Gene 1992; 112: 29–35. W. M. Barnes.

Crystal structures of open and closed forms of binary and ternary complexes of the large fragment of Thermus aquaticus DNA polymerase I: structural basis for nucleotide incorporation. 
EMBO J. 1998; 17: 7514–7525. Y. Li, S. Korolev and G. Waksman.

How can I optimize the PCR reaction conditions?

1. The annealing temperature can usually be optimized. Try a temperature gradient and determine the best annealing temperature, which yields in the cleanest product.

2. Add a gradual amount of betaine 0-1 M or DMSO 0-7.5% to the reaction mix and select for the cleanest product and the highest yield.

3. Try to shorten the extension and annealing time. Too long and too many cycles may lead to over-amplification and side-products.


img-contactContact & Support

img-accordionContact for support

img-accordionNeed project consulting?

img-accordionPublish and Get A Free Kit

 

img-downloadDownload

img-accordionMSDS (Material safety data sheet)

img-accordionProduct manual

img-accordionOrder form

 

 

Comments are closed.