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. KTQÂ DNA polymeraseÂ can also be used for realtime cycling, when adding a suitable realtime dye.
|#4001 S||KTQ DNA polymerase||400 U, 5U/Âµl, 80 Âµl||Â â‚¬ 45,-|
|#4001Â M||KTQ DNA polymerase||1000 U, 5U/Âµl, 200 Âµl||Â â‚¬ 105,-|
|#4001Â L||KTQ DNA polymerase||4000 U, 5U/Âµl, 2x 400 Âµl||Â â‚¬ 400,-|
Bulk offer: Should you require larger amounts (> 10.000 U), significant discounts are available. Please contact us to receive an individual quotation.
KTQ: Extreme thermostability
Comparison ofÂ 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 â€“ 97Â° C for one hour. PCR products were subsequently analysed on a 2.5% agarose gel.
|Primer forward (10 ÂµM)*||1 Âµl||0.2 ÂµM (0.05-1 ÂµM)|
|Primer reserve(10 ÂµM)*||1 Âµl||0.2 ÂµM (0.05-1 ÂµM)|
|dNTPs (2 mM)||5 Âµl||200 ÂµM|
|10 x KTQ buffer||5 Âµl||1x|
|KTQ DNA polymerase 5 U/Âµl||0.25 Âµl||1.25 U / reaction|
|Template/Sample extract**||x Âµl|
|Nuclease-free water||up to 50Âµl 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 Âµl (gnomic DNA) or 1 ng - 1 pg (plasmid/viral DNA).
- 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
|Initial denaturation||95Â°C||2 min|
|Annealing*||54-72Â°C||30 secÂ Â Â (25-40 cycles)|
|Extension||72Â Â°C||2Â min / 1000 bp|
*Typically, the annealing temperature is about 3-5Â°C below the calculated melting temperature of the primers used.Â
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 analyzedÂ byÂ 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 email@example.com for the lot-specific concentration.
No contamination has been detected in standard test reactions.
This product is shipped on cool packs. Please store the product upon arrival at -20Â°C.
Composition / Content of the Product
KTQ DNA polymerase is supplied as a 5 U/Âµl solution containing glycerol. It comes together with anÂ optimized 10x reaction buffer.
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: 29â€“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.Â
EMBO J. 1998; 17: 7514â€“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.
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