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Breast Cancer Research
Published in: Breast Cancer Research 1/2017

Open Access 01-12-2017 | Research Article

Nanopore sequencing of full-length BRCA1 mRNA transcripts reveals co-occurrence of known exon skipping events

Authors: Lucy C. de Jong, Simone Cree, Vanessa Lattimore, George A. R. Wiggins, Amanda B. Spurdle, Allison Miller, Martin A. Kennedy, Logan C. Walker, kConFab Investigators

Published in: Breast Cancer Research | Issue 1/2017

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Abstract

Background

Laboratory assays evaluating the effect of DNA sequence variants on BRCA1 mRNA splicing may contribute to classification by providing molecular evidence. However, our knowledge of normal and aberrant BRCA1 splicing events to date has been limited to data derived from assays targeting partial transcript sequences. This study explored the utility of nanopore sequencing to examine whole BRCA1 mRNA transcripts and to provide accurate categorisation of in-frame and out-of-frame splicing events.

Methods

The exon structure of BRCA1 transcripts from a previously studied control lymphoblastoid cell line were assessed using MinION nanopore sequencing of long-range reverse transcriptase-PCR amplicons.

Results

Our study identified and characterised 32 complete BRCA1 isoforms, including 18 novel isoforms which showed skipping of multiple contiguous and/or non-contiguous exons. Furthermore, we show that known BRCA1 exon skipping events, such as Δ(9,10) and Δ21, can co-occur in a single transcript, with some isoforms containing four or more alternative splice junctions. Fourteen novel isoforms were formed entirely from a combination of previously identified alternative splice junctions, suggesting that the total number of BRCA1 isoforms might be lower than the number of splicing events reported previously.

Conclusions

Our results highlight complexity in BRCA1 transcript structure that has not been described previously. This finding has key implications for predicting the translation frame of splicing transcripts, important for interpreting the clinical significance of spliceogenic variants. Future research is warranted to quantitatively assess full-length BRCA1 transcript levels, and to assess the application of nanopore sequencing for routine evaluation of potential spliceogenic variants.
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Metadata
Title
Nanopore sequencing of full-length BRCA1 mRNA transcripts reveals co-occurrence of known exon skipping events
Authors
Lucy C. de Jong
Simone Cree
Vanessa Lattimore
George A. R. Wiggins
Amanda B. Spurdle
Allison Miller
Martin A. Kennedy
Logan C. Walker
kConFab Investigators
Publication date
01-12-2017
Publisher
BioMed Central
Published in
Breast Cancer Research / Issue 1/2017
Electronic ISSN: 1465-542X
DOI
https://doi.org/10.1186/s13058-017-0919-1

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