Nondestructive enzymatic deamination enables single-molecule long-read amplicon sequencing for the determination of 5-methylcytosine and 5-hydroxymethylcytosine at single-base resolution.

The predominant methodology for DNA methylation analysis relies on the chemical deamination by sodium bisulfite of unmodified cytosine to uracil to permit the differential readout of methylated cytosines. Bisulfite treatment damages the DNA, leading to fragmentation and loss of long-range methylation information. To overcome this limitation of bisulfite-treated DNA, we applied a new enzymatic deamination approach, termed enzymatic methyl-seq (EM-seq), to long-range sequencing technologies.

RADAR-seq: A RAre DAmage and Repair sequencing method for detecting DNA damage on a genome-wide scale.

RAre DAmage and Repair sequencing (RADAR-seq) is a highly adaptable sequencing method that enables the identification and detection of rare DNA damage events for a wide variety of DNA lesions at single-molecule resolution on a genome-wide scale. In RADAR-seq, DNA lesions are replaced with a patch of modified bases that can be directly detected by Pacific Biosciences Single Molecule Real-Time (SMRT) sequencing. RADAR-seq enables dynamic detection over a wide range of DNA damage frequencies, including low physiological levels.

Solid-phase enzyme catalysis of DNA end repair and 3' A-tailing reduces GC-bias in next-generation sequencing of human genomic DNA.

The use of next-generation sequencing (NGS) has been instrumental in advancing biological research and clinical diagnostics. To fully utilize the power of NGS, complete, uniform coverage of the entire genome is required. In this study, we identified the primary sources of bias observed in sequence coverage across AT-rich regions of the human genome with existing amplification-free DNA library preparation methods.

Response to Comment on "DNA damage is a pervasive cause of sequencing errors, directly confounding variant identification".

Following the Comment of Stewart , we repeated our analysis on sequencing runs from The Cancer Genome Atlas (TCGA) using their suggested parameters. We found signs of oxidative damage in all sequence contexts and irrespective of the sequencing date, reaffirming that DNA damage affects mutation-calling pipelines in their ability to accurately identify somatic variations.

DNA damage is a pervasive cause of sequencing errors, directly confounding variant identification.

Mutations in somatic cells generate a heterogeneous genomic population and may result in serious medical conditions. Although cancer is typically associated with somatic variations, advances in DNA sequencing indicate that cell-specific variants affect a number of phenotypes and pathologies. Here, we show that mutagenic damage accounts for the majority of the erroneous identification of variants with low to moderate (1 to 5%) frequency.

Discovering novel cis-regulatory motifs using functional networks.

We combined functional information such as protein-protein interactions or metabolic networks with genome information in Saccharomyces cerevisiae to predict cis-regulatory motifs in the upstream region of genes. We developed a new scoring metric combining these two information sources and used this metric in motif discovery. To estimate the statistical significance of this metric, we used brute-force randomization, which shows a consistent well-behaved trend.