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Enzyme Finder NEBcutter NEBuffer Chart Double Digest Finder Isoschizomers DNA Sequences and Maps REBASE

Home > Products > Guidelines for Using Phusion Flash PCR Master Mix Guidelines for Using Phusion Flash PCR Master Mix Protocol Basic reaction conditions for PCR amplifications Carefully mix and spin down the Phusion Flash PCR Master Mix tube before opening to improve recovery. The PCR setup can be performed at room temperature. Pipetting instructions Component 50 µl reaction 20 µl reaction Final Conc. H2O add to 50 µl add to 20 µl   2x Phusion Flash PCR Master Mix 25 µl 10 µl 1X Primer A x µl x µl 0.5 µM Primer B x µl x µl 0.5 µM Template DNA x µl xµl   Cycling conditions: Cycling instructions Cycle step 2-step protocol 3-step protocol Cycles Temp. Time Temp. Time Initial denaturation 98°C 10 s 98°C 10 s 1 Denaturation Annealing Extension 98°C - 72°C 0 or 1 s - 15 s/kb 98°C X°C 72°C 0 or 1 s 5 s 15 s/kb 30 Final extension   72°C 4°C 1 min hold 70°C 4°C 1 min hold 1