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Full-length mRNA sequencing reveals principles of poly(A) tail length control

By Ivano Legnini, Jonathan Alles, Nikos Karaiskos, Salah Ayoub, Nikolaus Rajewsky

Posted 11 Feb 2019
bioRxiv DOI: 10.1101/547034 (published DOI: 10.1038/s41592-019-0503-y)

Although mRNAs are key molecules for understanding life, there exists no method to determine the full-length sequence of endogenous mRNAs including their poly(A) tails. Moreover, although poly(A) tails can be modified in functionally important ways, there also exists no method to accurately sequence them. Here, we present FLAM-seq, a rapid and simple method for high-quality sequencing of entire mRNAs. We report a cDNA library preparation method coupled to single-molecule sequencing to perform FLAM-seq. Using human cell lines, brain organoids, and C. elegans we show that FLAM-seq delivers high-quality full-length mRNA sequences for thousands of different genes per sample. We find that (a) 3' UTR length is correlated with poly(A) tail length, (b) alternative polyadenylation sites and alternative promoters for the same gene are linked to different tail lengths, (c) tails contain a significant number of cytosines. Thus, we provide a widely useful method and fundamental insights into poly(A) tail regulation.

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