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bioRxiv · 10.1101/149740

Mutational sequencing for accurate count and long-range assembly

Kumar, V.·Rosenbaum, J.·Wang, Z.·Forcier, T.·Ronemus, M.·Wigler, M.·Levy, D.
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Abstract

We introduce a new protocol, mutational sequencing or muSeq, which randomly deaminates unmethylated cytosines at a fixed and tunable rate. The muSeq protocol marks each initial template molecule with a unique mutation signature that is present in every copy of the template, and in every fragmented copy of a copy. In the sequenced read data, this signature is observed as a unique pattern of C-to-T or G-to-A nucleotide conversions. Clustering reads with the same conversion pattern enables accurate count and long-range assembly of initial template molecules from short-read sequence data. We explore count and low-error sequencing by profiling a 135,000 fragment PstI representation, demonstrating that muSeq improves copy number inference and significantly reduces sporadic sequencer error. We explore long-range assembly in the context of cDNA, generating contiguous transcript clusters greater than 3,000 bp in length. The muSeq assemblies reveal transcriptional diversity not observable from short-read data alone.

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Kumar, V., Rosenbaum, J., Wang, Z., Forcier, T., Ronemus, M., Wigler, M., Levy, D.. 2017-06-13. Mutational sequencing for accurate count and long-range assembly. https://doi.org/10.1101/149740

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