Protocol Prediction and Validation of Native and Engineered Cas9 Guide Sequences

This article describes a protocol for predicting and validating native and engineered Cas9 guide sequences. The protocol includes the following steps: (1) Identify the PAM sequence of the target gene; (2) Design the guide RNA sequence; (3) Synthesize the guide RNA; (4) Transfect the target cells; (5) Assess the editing efficiency.