DNA conformation may deviate from the classical B-form in ∼13% of the human genome. Non-B DNA regulates many cellular processes; however, its effects on DNA polymerization speed and accuracy have not been investigated genome-wide. Such an inquiry is critical for understanding neurological diseases and cancer genome instability. Here, we present the first simultaneous examination of DNA polymerization kinetics and errors in the human genome sequenced with Single-Molecule Real-Time (SMRT) technology. We show that polymerization speed differs between non-B and B-DNA: It decelerates at G-quadruplexes and fluctuates periodically at disease-causing tandem repeats. Analyzing polymerization kinetics profiles, we predict and validate experimentally non-B DNA formation for a novel motif. We demonstrate that several non-B motifs affect sequencing errors (e.g., G-quadruplexes increase error rates), and that sequencing errors are positively associated with polymerase slowdown. Finally, we show that highly divergent G4 motifs have pronounced polymerization slowdown and high sequencing error rates, suggesting similar mechanisms for sequencing errors and germline mutations.

Bioinformatics and Genomics Graduate Program Penn State University University Park Pennsylvania 16802 USA

Department of Biology Penn State University University Park Pennsylvania 16802 USA

Department of Biophysics of Nucleic Acids Institute of Biophysics of the Czech Academy of Sciences Královopolská 135 612 65 Brno Czech Republic

Department of Pathology Penn State University College of Medicine Hershey Pennsylvania 17033 USA

Department of Plant Developmental Genetics Institute of Biophysics of the Czech Academy of Sciences Královopolská 135 612 65 Brno Czech Republic

Department of Statistics Penn State University University Park Pennsylvania 16802 USA

Sant'Anna School of Advanced Studies 56127 Pisa Italy

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Accurate sequencing of DNA motifs able to form alternative (non-B) structures

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