Illumina sequencing of PANCE selection phages encoding TadA-8e deaminase
Plasmid phage DNA (1 ng) from each PANCE selection round was amplified using Q5 High-Fidelity 2 Master Mix (NEB) and primer pairs IllSeq_ABE_i5_F14 / IllSeq_ABE_i7_R14 containing i5/i7 overhangs and 47 nt random bases (N) for sequence diversity. A second PCR was performed using NEBNext Multiplex Oligos for Illumina to add full-length adapters and indexes. PCR products were purified using the Monarch Gel Extraction Kit (NEB) and quantified by Qubit High Sensitivity DNA Assay (Thermo Fisher). Equimolar libraries (4 nM each) were pooled, supplemented with 10% PhiX Control (4 nM), and sequenced on an Illumina MiSeq using the MiSeq Reagent Kit v3 (600 cycles, MS-102-3003) following the manufacturers protocol. V
SUB15721810
R5C_S15_L001_R1_001