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Amplicon Sequencing using the Long-Read Sequencing Technologies
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Long-Read Sequencing - A Powerful Tool in Viral Transcriptome Research.

Zsolt Boldogkői1, Norbert Moldován1, Zsolt Balázs1

  • 1Department of Medical Biology, Faculty of Medicine, University of Szeged, Szeged, Hungary.

Trends in Microbiology
|March 3, 2019
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Long-read sequencing (LRS) offers superior full-length RNA analysis and complex genome assembly. This technology reveals novel viral transcripts and intricate transcriptional overlaps, advancing our understanding of viral gene expression.

Keywords:
DNA replicationPacific BiosciencesRNA-Seqnanopore sequencingnoncoding RNAreplication originsplicing

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Area of Science:

  • Genomics and Transcriptomics
  • Virology
  • Molecular Biology

Background:

  • Long-read sequencing (LRS) technologies, such as Pacific Biosciences and Oxford Nanopore, are gaining prominence.
  • LRS excels in de novo assembly, resolving complex genomic regions, and characterizing full-length RNA molecules.
  • Despite lower coverage and higher error rates than short-read sequencing, LRS offers unique advantages for transcript analysis.

Purpose of the Study:

  • To highlight the utility of LRS in viral transcriptome analysis.
  • To demonstrate LRS's capability in identifying novel viral transcripts and complex transcriptional patterns.
  • To showcase LRS as an effective tool for studying small, compact viral genomes.

Main Methods:

  • Application of long-read sequencing technologies (Pacific Biosciences and Oxford Nanopore).
  • Analysis of viral transcriptomes to identify full-length RNA molecules.
  • Comparative analysis with short-read sequencing to underscore LRS advantages.

Main Results:

  • LRS identified a greater number of previously unknown viral transcripts compared to traditional methods.
  • Complex networks of overlapping transcripts were revealed in the studied viral genomes.
  • LRS demonstrated superiority in resolving transcript isoforms, including alternative splicing and polycistronic RNAs.

Conclusions:

  • Long-read sequencing is a powerful tool for comprehensive viral transcriptome analysis.
  • LRS significantly expands the discovery of viral genetic elements and regulatory mechanisms.
  • The application of LRS to viral studies is crucial for uncovering intricate transcriptional landscapes.