HOT FIREPol® Blend Master Mix Ready to Load with 10 mM MgCl2

A 5x-concentrated ready-to-use PCR master mix from Solis BioDyne supplies all reagents required for PCR except template, primers and water. The blend combines HOT FIREPol® DNA polymerase with a proofreading polymerase, providing both 5' flap endonuclease and 3'→5' proofreading activity. The manufacturer states that the mix shows increased fidelity, up to five-fold compared to HOT FIREPol®.

The formulation is supplied with 10 mM MgCl2 (delivering 2 mM MgCl2 in a 1x reaction) and 1 mM of each dNTP (200 µM of each dATP, dCTP, dGTP and dTTP at 1x), together with BSA. Two tracking dyes are included for monitoring electrophoresis progress: blue dye migrates equivalent to a 3.5–4.5 kb DNA fragment and yellow dye equivalent to 35–45 bp. A density-enhancing compound allows PCR reactions to be loaded directly onto agarose gels. The supplied mix is a green liquid at pH 7.0–9.0 and is not classified as hazardous under CLP criteria.

Generated PCR products are compatible with blunt-end and TA cloning. The mix is recommended for hot-start PCR applications visualised by agarose gel electrophoresis and ethidium bromide staining, and is not advised in workflows that rely on spectrophotometric absorbance or fluorescence readings. Suggested cycling conditions include an initial activation at 95°C for 12–15 minutes, then 25–30 cycles of denaturation at 95°C for 15–20 s, annealing at 54–66°C for 30–60 s and extension at 72°C for 20 s–4 min (a guideline of 1 min/kb is suggested), followed by a final extension at 72°C for 5–10 min. Reaction setup can be performed at room temperature. Routine storage is at -18°C to -28°C; shipping or temporary storage for up to one month at room temperature, or up to six months at 2–8°C, is reported to have no detrimental effect on quality. The product is manufactured by Solis BioDyne under ISO 9001 and ISO 13485 quality management systems and is supplied for research use only.