Annotation of full-length transcripts including alternative splicing from 19 chicken tissues using Oxford Nanopore long-read sequencing

GUAN, D.; GOSZCZYNSKI, D. E.; ZHOU, H.; ISLAS-TREJO, A. D.; ROSS, P. J.; HALSTEAD, M. M.; CHENG, H. H.

Conferencia; 2021 International Society for Animal Genetics Virtual Conference; 2021

The diversity of alternative splicing is one of major factors affecting phenotypic variability, while a comprehensive annotation of transcript isoforms across tissues has not been fully characterized. In this study, we utilized Oxford Nanopore Technology (ONT) with a large-scale multiplexing strategy, aiming to annotate full-length transcripts and alternative splicing in diverse chicken tissues derived from 68 biological samples, comprising 19 tissues from adult males and females of a F1 cross of line 6 ×line 7. The ONT cDNA RNA sequencing from a single ONT flow cell resulted in more than 19.4 million uniquely mapping full-length reads with mean read length of 648 bases, and average quality of 18.2. Using StringTie computational pipeline, we annotated 79,885 transcripts at 54,590 genetic loci, representing ~1.5 transcripts per locus. The sensitivity and precision of our annotation were 38.8% and 14.5% higher at the transcript level, and 57.7% and 17.5% higher at the locus level, respectively. For the annotated transcripts, there were 48,464 multi-exon and 31,421 single-exon transcripts. Compared with the Ensembl database (GRCg6a version 102), we annotated 2.7- and 3.3-fold more transcripts and gene loci, respectively. Among them, 14.6% (11,624) can fully match to the reference, and 35.4% are partially matched to known genes, and 39,985 annotated transcripts had unknown match. Further analysis will identify tissue-specific novel isoforms and their respective biological functions in chickens. In summary, this comprehensive annotation of transcript isoforms across diverse tissues has made significant contribution on the effort in functionally annotating chicken genome and provides novel knowledge in connecting genotype to phenotype in livestock species.