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(A) Cas9 cleavage pioneer assay was started by mixing radiolabeled DNA duplex and Mg2+ simultaneously to Cas9.gRNA, and the reaction was stopped at various time points. The scheme is shown above the gel image. (B) The plot of product formation as a function of time was fit by the double-exponential equation to obtain the cleavage rates for the initial fast phase and slow phase. Therefore, the reaction is biphasic. This figure is adapted from Gong et al. 2018.
