PacBio Iso-Seq Improves the Rainbow Trout Genome Annotation and Identifies Alternative Splicing Associated With Economically Important Phenotypes

Abstract

Rainbow trout is an important model organism that has received concerted international efforts to study the transcriptome. For this purpose, short-read sequencing has been primarily used over the past decade. However, these sequences are too short of resolving the transcriptome complexity. This study reported a first full-length transcriptome assembly of the rainbow trout using single-molecule long-read isoform sequencing (Iso-Seq). Extensive computational approaches were used to refine and validate the reconstructed transcriptome. The study identified 10,640 high-confidence transcripts not previously annotated, in addition to 1,479 isoforms not mapped to the current Swanson reference genome. Most of the identified lncRNAs were non-coding variants of coding transcripts. The majority of genes had multiple transcript isoforms (average ∼3 isoforms/locus). Intron retention (IR) and exon skipping (ES) accounted for 56% of alternative splicing (AS) events. Iso-Seq improved the reference genome annotation, which allowed identification of characteristic AS associated with fish growth, muscle accretion, disease resistance, stress response, and fish migration. For instance, an ES in GVIN1 gene existed in fish susceptible to bacterial cold-water disease (BCWD). Besides, under five stress conditions, there was a commonly regulated exon in prolyl 4-hydroxylase subunit alpha-2 (P4HA2) gene. The reconstructed gene models and their posttranscriptional processing in rainbow trout provide invaluable resources that could be further used for future genetics and genomics studies. Additionally, the study identified characteristic transcription events associated with economically important phenotypes, which could be applied in selective breeding.