FAQ: Why do I observe incomplete digestion?

  • Incomplete digestion may be caused by an incorrect ratio of Cas9 Nuclease, sgRNA, and target site. For complete digestion we recommend a 10:10:1 molar ratio of Cas9 Nuclease:sgRNA:target site. 
  • Incomplete digestion may be caused by a suboptimal sequence of the sgRNA. Verify the sequence and design of the sgRNA transcription template.
  • Incomplete digestion may be caused by poor quality sgRNA. Verify the integrity of the sgRNA by gel electrophoresis.
  • Incomplete digestion may be caused by using suboptimal buffer. Please use the NEBuffer 3.1 included with the enzyme.