Amplicon sequencig on all even wells in the baseline DNA-based CRISPR library
In brief, the baseline protocol entailed a reconditioning PCR with an initial round of PCR using universal primers for 30 cycles, followed by a 1:25 product dilution, and subsequent amplification with reduced cycles (10 cycles). Commercial sequencing was carried out by Novogene, utilizing paired-end, 150 bp, PCR-free library preparation using the NEBNext Ultra II DNA Library Prep Kit (NEB, catalog no. E7645) to achieve a deep coverage of 250x.