Figure 1. Outline of 3PC protocol. Total RNA is chemically fragmented and purified using oligo(dT) magnetic beads. Poly(A)-containing fragments are used as substrates for reverse transcription, and the resulting cDNAs are circularized and used as templates for PCR amplification. Poly(A) and poly(T) sequences are indicated. “R” represents the random sequence of the unique molecular identifier, and “B” a multiplexing barcode. Sequences from the RT primer that are used for PCR amplification are represented in cyan and green. See Materials and Methods for more details.