Before starting the protocol, the quality of the gDNA needs to be assessed. We recommend using unsheared, high-molecular-weight DNA at ≥210 ng/µl by Qubit and stored in TE (pH 8.0) or similar, or nuclease-free water.
During the CRISP/Cas9 sequencing protocol, the target regions will be available for sequencing adapter ligation but the background DNA is still present, therefore quantification of the sample at different stages won’t be a good indication of how well the library prep is going.
For positive control, we recommend the validated crRNA probes for the human HTT gene. The probe sequences are provided here.
Please note that they are designed against the human genome. They can be combined in the initial RNP complex with your target probes and will act as an in-run control.