Capsid 1AA Insertion Library Passage 2 Replicate A
1 g of sample RNA was used in a ProtoScript II reverse transcription reaction (New EnglandBiolabs, E6560L) to produce cDNA by spiking in a gene-specific primerat the start of the 2Agene or in the 3 UTR to allow the amplification of the P1 and P2-P3 regions of the genomerespectively. Then, the cDNA or plasmid DNA was amplified to generate amplions using the Q5High-Fidelity DNA Polymerase in four independent PCR reactions for 25 cycles to minimizePCR-derived errors. The amplicons were gel-purified using the Monarch DNA Gel Extraction Kitand used as inputs for Illumina sequencing library preparation. Illumina sequencing librarieswere prepared using the Twist Biosciences Enzymatic Fragmentation 2.0 kit with universaladapters (Twist Biosciences, 104207) targeting fragment sizes inthe range 180-220 bp. Illuminalibraries were run on a MiSeq device using the MiSeq reagent kit v2, 300-cycles (Illumina,MS-102-2002).