Single-stranded DNA library preparation uncovers the origin and diversity of ultrashort cell-free DNA in plasma

Circulating cell-free DNA (cfDNA) is emerging as a powerful monitoring tool in cancer, pregnancy and organ transplantation. Nucleosomal DNA, the predominant form of plasma cfDNA, can be adapted for sequencing via ligation of double-stranded DNA (dsDNA) adapters. dsDNA library preparations, however,...

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Veröffentlicht in:Scientific reports 2016-06, Vol.6 (1), p.27859-27859, Article 27859
Hauptverfasser: Burnham, Philip, Kim, Min Seong, Agbor-Enoh, Sean, Luikart, Helen, Valantine, Hannah A., Khush, Kiran K., De Vlaminck, Iwijn
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Sprache:eng
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Zusammenfassung:Circulating cell-free DNA (cfDNA) is emerging as a powerful monitoring tool in cancer, pregnancy and organ transplantation. Nucleosomal DNA, the predominant form of plasma cfDNA, can be adapted for sequencing via ligation of double-stranded DNA (dsDNA) adapters. dsDNA library preparations, however, are insensitive to ultrashort, degraded cfDNA. Drawing inspiration from advances in paleogenomics, we have applied a single-stranded DNA (ssDNA) library preparation method to sequencing of cfDNA in the plasma of lung transplant recipients (40 samples, six patients). We found that ssDNA library preparation yields a greater portion of sub-100 bp nuclear genomic cfDNA ( p 10 −5 , Mann-Whitney U Test), and an increased relative abundance of mitochondrial (10.7x, p 10 −5 ) and microbial cfDNA (71.3x, p 10 −5 ). The higher yield of microbial sequences from this method increases the sensitivity of cfDNA-based monitoring for infections following transplantation. We detail the fragmentation pattern of mitochondrial, nuclear genomic and microbial cfDNA over a broad fragment length range. We report the observation of donor-specific mitochondrial cfDNA in the circulation of lung transplant recipients. A ssDNA library preparation method provides a more informative window into understudied forms of cfDNA, including mitochondrial and microbial derived cfDNA and short nuclear genomic cfDNA, while retaining information provided by standard dsDNA library preparation methods.
ISSN:2045-2322
2045-2322
DOI:10.1038/srep27859