Quick Tips - Why do I have so many wildtype colonies?

This video explains why you might have a large number of wildtype colonies following site-directed mutagenesis and transformation.

Script

Customers often ask, “Why is the background so high? Why do I have so many wildtype colonies?”

Wildtype colonies result from template DNA that is carried over from the PCR into the final transformation step. To eliminate template DNA, first, be sure that the PCR reaction has worked efficiently. Then, use only 1ul of the PCR reaction in the KLD reaction. This will eliminate plasmid DNA and reduce the number of background colonies.

If you have other questions you can always contact us at info@neb.com.

Related Videos

  • TroubleshootingTipsQ5SiteDirectedMutagenisis_720
    Troubleshooting tips for Q5® Site-Directed Mutagenesis Kit
  • NEBTV_DNA-Assembly-Cloning-and-Mutagenesis-Kits_1920
    Quick Tips - Can I make multiple mutations with the Q5® site-directed mutagenesis kit?
  • OverviewOfQ5SDMKit_720
    Overview of the Q5® Site-Directed Mutagenesis Kit
Visit NEB’s Video Library
Loading Spinner