Full-length alternative transcript isoform analysis with nanopore sequencing

Our group aims to understand the mechanisms of alternative RNA splicing regulation and splicing dysregulation in cancer. Short-read, high-throughput cDNA sequencing (RNA-Seq) has revolutionized our ability to profile RNA splicing; however, this approach cannot capture the full complexity of RNA transcripts. First, “RNA-Seq” should, more appropriately, be called cDNA-Seqâ€"it is not sequencing RNA directly. Second, short-reads limit our ability to accurately identify and quantify full-length RNA isoforms. For a more comprehensive characterization of alternative transcript isoform expression, we have been developing computational approaches to analyze long-read nanopore sequencing data. I will present a study to identify differentially expressed isoforms from nanopore cDNA sequencing of isogenic cell lines with and without a mutation in U2AF1, which is a recurrently mutated splicing factor in cancer. I will also present our analysis of native RNA sequencing of the GM12878 cell line, as part of the Nanopore RNA Consortium. Utilizing the full benefit of directly sequencing full-length RNA transcripts, we identified alternative transcript isoforms and their association with allele expression, RNA modifications, and poly(A) tail length.

Speaker: Angela Brooks, UC Santa Cruz