HOT FIREPol® DNA Polymerase
Description
HOT FIREPol® is a chemically modified FIREPol® DNA Polymerase.
HOT FIREPol® is inactive at room temperature and is activated by a 15 min incubation step at 95°C. This enables hot-start PCR and minimizes mispriming and extension from non-specifically annealed primers and primer-dimers. Recommended for routine applications (fragment up to 5 kb).
Possesses 5′→3′ polymerase and 5′→3′ exonuclease activity, as well as a non-template-dependent terminal transferase activity, but lacks a 3′→5′ exonuclease (proofreading) activity making the generated product suitable for TA-cloning.
The fidelity of HOT FIREPol® is similar to a regular Taq DNA Polymerase (error rate per nucleotide app. 2.5 x10-5).
Properties
Concentration: 5 U/µl
Hot-start: yes, activated by a 15 min incubation step at 95°C
Error rate per nucleotide per cycle is app. 2.5 x 10-5
Accuracy is app. 4 x 104
Estimated half-life at 95ºC is 1.5 hours.
Storage and Dilution Buffer: 50% glycerol (v/v), 20 mM Tris-HCl pH
Source: Purified from an E.coli strain that carries an overproducing plasmid containing a modified gene of Thermus aquaticus DNA Polymerase.
Reagents
HOT FIREPol® DNA Polymerase (5 U/µl) in 20 mM Tris-HCl pH 8.7 at 25ºC, 100 mM KCl, 0.1 mM EDTA, 50% glycerol (v/v), and stabilizers.
HOT FIREPol® 10x Buffer B1 (without Mg2+ and detergent): 0.7 M Tris-HCl, 0.175 M (NH4)2SO4.
HOT FIREPol® 10x Buffer B2 (without Mg2+): 0.7 M Tris-HCl, 0.175 M (NH4)2SO4, 0.2% w/v Tween-20. HOT FIREPol® 10x Buffer B2 contains non-ionic detergent suppressing inhibitory effects of the trace of DNA extraction buffers and enhancing PCR yield and efficiency.
25 mM MgCl2
GC-rich enhancer is an additive that facilitates the amplification of difficult templates (e.g. GC-rich DNA templates). This solution should be used at a defined final concentration (1x, 2x or 3x solution). GC-rich enhancer is NOT a reaction buffer and should be used ONLY IF non-specific amplification occurs.
