HOT FIREPol® EvaGreen® HRM Mix (no ROX)

This 5x concentrated ready-to-use solution from Solis BioDyne is formulated for High Resolution Melt (HRM) Analysis. Designated HOT FIREPol® EvaGreen® HRM Mix (no ROX), it incorporates EvaGreen® dye and provides all components necessary to perform qPCR and HRM Analysis: HOT FIREPol® DNA Polymerase, ultrapure dNTPs, MgCl2 and EvaGreen® dye. The user adds only water, template and primers.

The mix contains a 5x EvaGreen® HRM buffer with 12.5 mM MgCl2 (yielding 2.5 mM MgCl2 in the 1x PCR solution). HOT FIREPol® DNA Polymerase is activated by a 12 min incubation at 95°C, which prevents extension of non-specifically annealed primers and primer-dimers formed during qPCR setup. A recommended 20 µl qPCR reaction uses 4 µl of the 5x mix with 80–250 nM each primer and variable template. The recommended cycling protocol includes an initial 95°C activation for 12 min, followed by 40 cycles of 95°C for 15 s, annealing at 60°–65°C for 20 s, and extension at 72°C for 20 s (extending annealing/extension to 30 s for templates longer than 150 bp). EvaGreen® is described as having much less PCR inhibition, being extremely stable, and non-mutagenic and non-cytotoxic, and is compatible with all common real-time PCR cyclers using standard SYBR® Green or FAM settings.

The product is stored routinely at -18°C to -28°C; shipping and temporary storage for up to 1 month at room temperature is stated to have no detrimental effects on quality. It is manufactured by Solis BioDyne in compliance with ISO 9001 and ISO 13485 certified Quality Management Systems. For research use only; not for use in diagnostic procedures.