Preparation and analysis of 5′ cap-adapted poly(A) strands. (A) Adaptation and library preparation workflow for poly(A)-selected RNA. (B) Representative ionic current trace for a cap-adapted full-length RNA read is shown for the thymidine phosphorylase gene (TYMP). The trace begins with ionic current associated with the ONT adapter (i). This is followed by a monotonic ionic current associated with the 3′ poly(A) tail (ii) and then a variable ionic current associated with the RNA transcript nucleotides (iii). The final segment is an ionic current signature characteristic of the 45 nt RNA cap-adapter (iv). (C) An approximately two second window centered on the ionic current associated with the poly(A) tail (ii). (D) An approximately one second window centered on the boundary between the ionic current associated with the 5′ end of the transcript (iii) and a characteristic adapter ionic current trace (iv).
