Distinguishing amplification artifacts from biologically derived somatic mutations in single-cell sequencing data

Linked-read analysis identifies mutations in single-cell DNA-sequencing data

Craig L. Bohrson, Alison R. Barton, Michael A. Lodato, Rachel E. Rodin, Lovelace J. Luquette, Vinay V. Viswanadham, Doga C. Gulhan, Isidro Cortés-Ciriano, Maxwell A. Sherman, Minseok Kwon,  Michael E. Coulter, Alon Galor, Christopher A. Walsh & Peter J. Park

Nature Genetics (Research Article)

Whole-genome sequencing of DNA from single cells has the potential to reshape our understanding of mutational heterogeneity in normal and diseased tissues. However, a major difficulty is distinguishing amplification artifacts from biologically derived somatic mutations. Here, we describe linked-read analysis (LiRA), a method that accurately identifies somatic single-nucleotide variants (sSNVs) by using read-level phasing with nearby germline heterozygous polymorphisms, thereby enabling the characterization of mutational signatures and estimation of somatic mutation rates in single cells.

Next-next generation tool for improving traditional chemotherapies

Direct Analysis of Incorporation of an Anticancer Drug into DNA at Single-Molecule Resolution

Identifying positions at which anticancer drug molecules incorporate into DNA is essential to define mechanisms underlying their activity, but current methodologies cannot yet achieve this. The thymidine fluorine substitution product trifluridine (FTD) is a DNA-damaging anticancer agent thought to incorporate into thymine positions in DNA. This mechanism, however, has not been directly confirmed. Here, we report a means to detect FTD in a single-stranded oligonucleotide using a method to distinguish single molecules by differences in electrical conductance. Entire sequences of 21-base single-stranded DNAs with and without incorporated drug were determined based on single-molecule conductances of the drug and four deoxynucleosides, the first direct observation of its kind. This methodology may foster rapid development of more effective anticancer drugs.

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