Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus

Wen Chun Liu, Chih Peng Lin, Chun Pei Cheng, Cheng Hsun Ho, Kuo Lun Lan, Ji Hong Cheng, Chia Jui Yen, Pin Nan Cheng, I. Chin Wu, I. Chen Li, Bill Chia Han Chang, Vincent S. Tseng, Yen Cheng Chiu, Ting Tsung Chang*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

Abstract

Background: Hepatitis B virus (HBV) quasispecies are crucial in the pathogenesis of chronic liver disease. Next-generation sequencing (NGS) is powerful for identifying viral quasispecies. To improve mapping quality and single nucleotide variant (SNV) calling accuracy in the NGS analysis of HBV, we compared different mapping references, including the sample-specific reference sequence, same genotype sequences and different genotype sequences, according to the sample. Methods: Real Illumina HBV datasets from 86 patients, and simulated datasets from 158 HBV strains in the GenBank database, were used to assess mapping quality. SNV calling accuracy was evaluated using different mapping references to align Real Illumina datasets from a single HBV clone. Results: Using the sample-specific reference sequence as a mapping reference produced the largest number of mappable reads and coverages. With a different genotype mapping reference, the consensus sequence derived from the Real Illumina datasets of the single HBV clone showed 21 false SNV callings in polymerase and surface genes, the regions most divergent between the mapping reference and this HBV clone. A ~6 % coverage of most of these false SNVs was yielded even with a same genotype mapping reference, but none with the sample-specific reference sequence. Conclusions: Using sample-specific reference sequences as a mapping reference in NGS analysis optimized mapping quality and the SNV calling accuracy for HBV quasispecies.

Original languageEnglish
Pages (from-to)147-157
Number of pages11
JournalHepatology International
Volume10
Issue number1
DOIs
StatePublished - 1 Jan 2016

Keywords

  • Alignment stage
  • Coverage
  • Divergence
  • Single nucleotide variants

Fingerprint Dive into the research topics of 'Aligning to the sample-specific reference sequence to optimize the accuracy of next-generation sequencing analysis for hepatitis B virus'. Together they form a unique fingerprint.

Cite this