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FAQ: Why do I observe incomplete binding with EnGen® Spy dCas9?

Incomplete binding may be due to the following factors:
  • Incorrect ratio of dCas9 to guide RNA, and target site. For complete binding we recommend a 10:10:1 or higher molar ratio (EnGen® Spy dCas9 : guide RNA : target site).
  • Suboptimal sequence of the guide RNA. Verify the sequence and design of the guide RNA.
  • Poor quality guide RNA. Verify the integrity of the guide RNA by gel electrophoresis.
  • Use of a suboptimal buffer. Please use NEBuffer 3.1, which is included with the enzyme.