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. Author manuscript; available in PMC: 2023 Jul 13.
Published in final edited form as: Nat Protoc. 2022 Nov 16;18(2):396–423. doi: 10.1038/s41596-022-00766-8

Extended Data Fig. 1. Sequence alignment for a representative PepSeq probe with amplification and sequencing primers.

Extended Data Fig. 1

Top: For DNA amplification, the forward or reverse primers bind to the PepSeq probe via the 19-nt constant regions added to either end of the DNA tag. The forward DNA amplification primer contains a T7 promoter, NEB untranslated region, start codon and TEV cleavage site sequences. The reverse DNA amplification primer contains an S6 tag and a CP1 annealing site. Bottom: For sequencing, the forward indexing primer contains a 12-nt randomer (N) and a 10-nt barcode sequence (B). The reverse indexing primer contains a separate 8-nt barcode (B). Both indexing primers bind to the DNA tags via the 19-nt constant regions. For the reverse primers, we are showing the reverse complement sequences to clearly indicate annealing regions. See Supplementary Table 1 for oligonucleotide sequences to order.