FAQ: Can I use Authenticase™ genome editing (CRISPR/Cas9, TALEN, ZFN) mismatch detection assays with unpurified PCR products?

Yes. For quick analysis, we recommend using 6 μl of the appropriate PCR reaction directly in a 20 µl Authenticase reaction when the following PCR reagents are used: Q5^® Hot Start High-Fidelity 2x Master Mix  (NEB #M0494) or OneTaq DNA Polymerase (NEB #M0480). Alternatively, one can use 1-12 μl of DNA processed by Exo-CIP™ Rapid PCR Cleanup Kit (NEB #E1050) directly in a 20 µl Authenticase reaction when the following PCR reagents are used: Q5 High-Fidelity DNA Polymerase (NEB #M0491) with Q5 Reaction Buffer (up to 6 µl) or OneTaq DNA Polymerase Reaction Buffer (up to 12 µl). Additional details are in the product manual. For a more accurate estimation of genomic editing efficiency, amplicons purified from Monarch^® PCR & DNA Cleanup kit (NEB #T1030) are recommended.