CLT-seq as a universal homopolymer-sequencing concept reveals poly(A) tail-tuned ncRNA regulation

To more understand the poly(A) tail-tuned regulation in eukaryotic cells, we describe central limit theorem (CLT)-managed RNA-seq (CLT-seq) as a universal homopolymer-sequencing concept that features unbiased RNA-seq and facilitates accurate homopolymer base-calling. We then performed transcriptome-wide poly(A)-tail profiling analysis using data obtained from oligo(dT)-primed RNA-seq of HEK293T and A549 cell lines. Comparative poly(A)-tail profiling analysis of RNA-seq data for different cells. The T12, T18, T30, T18V, T18VN, and T18VNN are adaptor-flanked oligo(dT) primers with anchor or without anchor base for poly(A) tail-specific reverse transcription. The sequence is as following: T12: 5'-TACGAGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTAGCGTAGCTTTTTTTTTTTT-3' T18: 5'-TACGAGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTAGCGTAGCTTTTTTTTTTTTTTTTTT-3' T30: 5'-TACGAGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTAGCGTAGCTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT-3' T18V: 5'-TACGAGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTAGCGTAGCTTTTTTTTTTTTTTTTTTV-3' T12VN: 5'-TACGAGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTAGCGTAGCTTTTTTTTTTTTTTTTTTVN-3' T18VNN: 5'-TACGAGATGTGACTGGAGTTCAGACGTGTGCTCTTCCGATCTAGCGTAGCTTTTTTTTTTTTTTTTTTVNN-3'