Tina O'Grady

Global transcript structure resolution of high gene density genomes through multi-platform data integration

Annotation of herpesvirus genomes has traditionally been undertaken through the detection of open reading frames and other genomic motifs, supplemented with sequencing of individual cDNAs. Second generation sequencing and high-density microarray studies have revealed vastly greater herpesvirus transcriptome complexity than is captured by existing annotation. The pervasive nature of overlapping transcription throughout herpesvirus genomes, however, poses substantial problems in resolving transcript structures using these methods alone. We present an approach that combines the unique attributes of Pacific Biosciences Iso-Seq long-read, Illumina short-read and deepCAGE (Cap Analysis of Gene Expression) sequencing to globally resolve polyadenylated isoform structures in replicating Epstein-Barr virus (EBV). Our method, Transcriptome Resolution through Integration of Multi-platform Data (TRIMD), identifies nearly 300 novel EBV transcripts, quadrupling the size of the annotated viral transcriptome. These findings illustrate an array of mechanisms through which EBV achieves functional diversity in its relatively small, compact genome including programmed alternative splicing (e.g. across the IR1 repeats), alternative promoter usage by LMP2 and other latency-associated transcripts, intergenic splicing at the BZLF2 locus, and antisense transcription and pervasive readthrough transcription throughout the genome.

Analysis of EBV Transcription Using High-Throughput RNA Sequencing.

High-throughput sequencing of RNA is used to analyze the transcriptomes of viruses and cells, providing information about transcript structure and abundance. A wide array of programs and pipelines has been created to manage and interpret the abundance of data generated from high-throughput RNA sequencing experiments. This protocol details the use of free and open-source programs to align RNA-Seq reads to a reference genome, visualize read coverage and splice junctions, estimate transcript abundance, and evaluate differential expression of transcripts in different conditions. Particular concerns related to EBV and viral transcriptomics are addressed and access to EBV reference files is provided.

Tumor microenvironment affects the composition of endothelial-derived extracellular vesicles: Impact in tumor progression

Book: ISEV2017 To cite this article: (2017) Abstract Book: ISEV2017, Journal of Extracellular Vesicles, 6:sup1, 1310414, DOI: 10.1080/20013078.2017.1310414 To link to this article: https://doi.org/10.1080/20013078.2017.1310414