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DCS Pol

DCSPol is a highly processive, thermostable DNA polymerase. Due to its genetic modifications DCSPol has an enhanced stability at room temperature with no activity loss for up to 1 month.

Source

Purified from an E.coli strain that carries an overproducing plasmid containing a modified gene of Thermus aquaticus DNA Polymerase.

Applications

  • Suited for a wide range of PCR assays
  • TA cloning

Reagents Provided

  • DCSPol DNA Polymerase
  • 10 x Reaction buffer B (Mg2+ free) 0.8 M Tris-HCl, 0.2 M (NH4)2SO4, 0.2% w/v Tween-20
  • 10 x Reaction buffer BD (Mg2+ and detergent free) 0.8 M Tris-HCl, 0.2 M (NH4)2SO4
  • 25 mM MgCl2
  • 10 x Solution S
    • Additive that facilitates amplification of difficult templates (e.g. GC-rich DNA templates). This solution should be used at a defined working concentration (1x, 2x or 3x solution). Solution S is not a reaction buffer and should be used only if non-specific amplifications occur.

Concentration

5 U/µl

Unit Definition

One unit is defined as the amount of enzyme required to catalyze the incorporation of 10 nmol of dNTPs into an acid-insoluble form in 30 minutes at 74ºC.

Storage and Dilution buffer

50% glycerol (v/v), 20 mM Tris-HCl pH 8.7 at 25ºC, 100 mM KCl, 0.1 mM EDTA and stabilizers.

Shipping and Storage conditions

Routine storage: -20ºC Shipping and temporary storage for up to 1 month at room temperature has no detrimental effects on the quality of DCSPol DNA Polymerase.

Quality Control

The enzyme is free of nicking and priming activities, exonucleases and non-specific endonucleases. SDS/PAGE - 95 kD band, >98% pure. Activity and stability tested via thermo-cycling. The error rate per nucleotide per cycle is ~ 2.5 x 10-5; the accuracy is ~ 4 x 104. Estimated half life at 95ºC is 1.5 hours.

Recommended PCR reaction mix

Component Volume Final conc.
DCSPol (5 U/µl) 0.4-1.0 µl 0.02-0.05 U/µl (2-5 U)
10 x Buffer B or BD 10 µl 1x
25 mM MgCl2 6-10 µl 1.5-2.5 mM
20 mM dNTP mix 1 µl 200 µM
Primer Forward (10 pmol/µl) 1-3 µl 0.1-0.3 µM
Primer Reverse (10 pmol/µl) 1-3 µl 0.1-0.3 µM
DNA template 5-20 µl 5-100 ng/µl
10 x Solution S Not for standard PCR 0, 10, 20 or 30 µl 1x, 2x or 3x
H2O PCR grade Up to 100 µl
Total 100 µl

Recommended PCR Cycles

Cycle step Temp. Time Cycles
Initial denaturation 95ºC 3-5 min 1
Denaturation 95ºC 30-60 s 26-35
Annealing 50-68ºC 30-60 s
Elongation 72ºC 1-4 min
Final elongation 72ºC 5-10 min 1
IMPORTANT: Annealing temperature should be 2-6ºC lower than the primer melting temperature. Elongation time should be ~1 min/1 kb.

Safety warnings and precautions

This product and its components should be handled only by persons trained in laboratory techniques. It is advisable to wear suitable protective clothing, such as laboratory overalls, gloves and safety glasses. Care should be taken to avoid contact with skin or eyes. In case of contact with skin or eyes, wash immediately with water. Some applications this product is used in may require a license which is not provided by the purchase of this product. Users should obtain the license if required.