FAQ: What causes an occasional smear in a "negative control" with no template present?

DNA polymerases with low Km values for DNA, especially those with proofreading exonuclease functions, can cause primer artifacts to form if the DNA polymerase cannot bind to its preferred substrate (a 3´ end of an annealed primer). This primer artifact possesses single-stranded and double-stranded regions, and can appear either as DNA barely migrating out of the gel well or as a smear originating at the gel well. OneTaq® (NEB #M0480), OneTaq® Hot Start (NEB #M0481), Q5® Hot Start (NEB# M0493), Phusion® Hot Start Flex (NEB #M0535) and Taq DNA Polymerases (NEB #M0267, NEB #M0273) can minimize this artifact.

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