Rxivist logo

Rxivist combines preprints from bioRxiv with data from Twitter to help you find the papers being discussed in your field. Currently indexing 57,910 bioRxiv papers from 266,458 authors.

PacBio library preparation using blunt-end adapter ligation produces significant artefactual fusion DNA sequences

By Paul M Griffith, Castle Raley, David Sun, Yongmei Zhao, Zhonghe Sun, Monika Mehta, Bao Tran, Xiaolin Wu

Posted 09 Jan 2018
bioRxiv DOI: 10.1101/245241

Pacific Biosciences (PacBio) RS II sequencer, utilizing Single-Molecule, Real-Time (SMRT) technology, has revolutionized next-generation sequencing by providing an accurate long-read platform. PacBio single-molecule long reads have been used to delineate complex spliceoforms, detect mutations in highly homologous sequences, identify mRNA chimeras and chromosomal translocations, accurately haplotype phasing over multiple kilobase distances and aid in assembly of genomes with complex structural variation. The PacBio protocol for preparation of sequencing templates employs blunt-end hairpin adapter ligation, which enables a short turnaround time for sequence production. However, we have found a significant portion of sequencing yield contains chimeric reads resulting from blunt-end ligation of multiple template molecules to each other prior to adapter ligation. These artefactual fusion DNA sequences pose a major challenge to analysis and can lead to false-positive detection of fusion events. We assessed the frequency of artefactual fusion when using blunt-end adapter ligation and compared it to an alternative method using A/T overhang adapter ligation. The A/T overhang adapter ligation method showed a vast improvement in limiting artefactual fusion events and is now our recommended procedure for adapter ligation during PacBio library preparation.

Download data

  • Downloaded 1,072 times
  • Download rankings, all-time:
    • Site-wide: 5,839 out of 57,910
    • In genetics: 481 out of 3,319
  • Year to date:
    • Site-wide: 8,554 out of 57,910
  • Since beginning of last month:
    • Site-wide: 10,543 out of 57,910

Altmetric data


Downloads over time

Distribution of downloads per paper, site-wide


Sign up for the Rxivist weekly newsletter! (Click here for more details.)


News