Long-read targeted sequencing uncovers clinicopathological associations for C9orf72-linked diseases

Mariely DeJesus-Hernandez, Ross A. Aleff, Jazmyne L. Jackson, Ni Cole A. Finch, Matthew C. Baker, Tania F. Gendron, Melissa E. Murray, Ian J. McLaughlin, John R. Harting, Neill R. Graff-Radford, Björn Oskarsson, David S. Knopman, Keith A. Josephs, Bradley F. Boeve, Ronald C. Petersen, John D. Fryer, Leonard Petrucelli, Dennis W. Dickson, Rosa Rademakers, Mark T.W. EbbertEric D Wieben, Marka Van Blitterswijk

Research output: Contribution to journalArticlepeer-review


To examine the length of a hexanucleotide expansion in C9orf72, which represents the most frequent genetic cause of frontotemporal lobar degeneration and motor neuron disease, we employed a targeted amplification-free long-read sequencing technology: No-Amp sequencing. In our cross-sectional study, we assessed cerebellar tissue from 28 well-characterized C9orf72 expansion carriers. We obtained 3507 on-target circular consensus sequencing reads, of which 814 bridged the C9orf72 repeat expansion (23%). Importantly, we observed a significant correlation between expansion sizes obtained using No-Amp sequencing and Southern blotting (P = 5.0 × 10-4). Interestingly, we also detected a significant survival advantage for individuals with smaller expansions (P = 0.004). Additionally, we uncovered that smaller expansions were significantly associated with higher levels of C9orf72 transcripts containing intron 1b (P = 0.003), poly(GP) proteins (P = 1.3 × 10- 5), and poly(GA) proteins (P = 0.005). Thorough examination of the composition of the expansion revealed that its GC content was extremely high (median: 100%) and that it was mainly composed of GGGGCC repeats (median: 96%), suggesting that expanded C9orf72 repeats are quite pure. Taken together, our findings demonstrate that No-Amp sequencing is a powerful tool that enables the discovery of relevant clinicopathological associations, highlighting the important role played by the cerebellar size of the expanded repeat in C9orf72-linked diseases.

Original languageEnglish (US)
Pages (from-to)1082-1088
Number of pages7
JournalBrain : a journal of neurology
Issue number4
StatePublished - May 7 2021


  • amyotrophic lateral sclerosis
  • C9orf72
  • frontotemporal lobar degeneration
  • long-read sequencing
  • motor neuron disease

ASJC Scopus subject areas

  • Clinical Neurology

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