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Defense: Documentation of high-confidence full-length human mRNA scaffolds based on single native RNA reads

Speaker Name: 
Logan Mulroney
Speaker Title: 
PhD Candidate
Speaker Organization: 
Biomolecular Engineering & Bioinformatics PhD
Start Time: 
Thursday, September 17, 2020 - 4:00pm
End Time: 
Thursday, September 17, 2020 - 5:00pm
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Abstract: Nanopore sequencing devices read individual RNA strands directly. This facilitates identification of exon linkages and nucleotide modifications, however using conventional methods the 5′ and 3′ ends of mature mRNA cannot be identified unambiguously. This is due in part to the architecture of the nanopore/enzyme-motor complex, and in part to RNA degradation in vivo and in vitro. In this study, we aimed to identify individual high-confidence full-length human mRNA isoform scaffolds among ~4 million nanopore poly(A)-selected RNA reads. First, we exchanged the biological 5′ m7G cap for a modified cap bearing a 45-nucleotide oligomer. This oligomer improved 5′ end sequencing and ensured identification of capped strands. Second, among these capped reads, we screened for 3′ ends consistent with documented polyadenylation sites. This gave 185,434 high-confidence mRNA scaffolds, including 4,262 that represented isoforms absent from GENCODE. Most of these had transcription start sites internal to longer, previously identified mRNA isoforms. Combined with orthogonal data, these mRNA scaffolds provide decisive evidence for full-length mRNA isoforms.

Event Type: 
Mark Akeson
Graduate Program: 
Biomolecular Engineering & Bioinformatics PhD