Multi-Platform Sequencing Approach Reveals a Novel Transcriptome Profile in Pseudorabies Virus

Area of Science:

• * Virology
• * Molecular Biology
• * Genomics

Background:

• * Third-generation sequencing technologies offer advanced capabilities for analyzing complex transcriptomes.
• * Previous studies on the pseudorabies virus (PRV) transcriptome were limited by earlier sequencing methods.
• * Understanding PRV's full transcriptional landscape is crucial for viral research.

Purpose of the Study:

• * To comprehensively analyze the PRV transcriptome using long-read sequencing.
• * To identify novel protein-coding genes, non-coding RNAs, and transcript isoforms.
• * To characterize transcript overlaps and their potential functional implications.

Main Methods:

• * Utilization of multiple long-read sequencing platforms: Oxford Nanopore MinION and PacBio RS-II.
• * Integration of data from Illumina HiScanSQ short-read sequencing.
• * Comparative analysis with previous short-read and long-read sequencing data for validation.

Main Results:

• * Discovery of 19 novel putative protein-coding genes, primarily 5' truncated forms.
• * Identification of 19 non-coding RNAs, including truncated transcripts and antisense RNAs.
• * Characterization of 3 complex transcripts, 50 distinct length isoforms, and 121 novel transcript overlaps.
• * Detection of two transcripts overlapping PRV replication origins.
• * In silico identification of 145 upstream open reading frames (ORFs).

Conclusions:

• * Third-generation sequencing significantly expands the known PRV transcriptome.
• * Novel findings include previously uncharacterized genes, RNAs, and complex transcriptional events.
• * This study provides a more complete view of PRV gene expression and regulation.