Library preparation and MiSeq sequencing for the genotyping-by-sequencing of the Huntington disease HTT exon one trinucleotide repeat and the quantification of somatic mosaicism [Protocol]

Ciosi, M. , Cumming, S. A., Alshammari, A. M., Symeonidi, E., Herzyk, P. , Mcguinness, D. , Galbraith, J., Hamilton, G. and Monckton, D. G. (2018) Library preparation and MiSeq sequencing for the genotyping-by-sequencing of the Huntington disease HTT exon one trinucleotide repeat and the quantification of somatic mosaicism [Protocol]. Other. Springer Nature. (doi: 10.21203/rs.2.1581/v2).

Abstract

Huntington disease (HD) is an autosomal dominant neurodegenerative disorder caused by the expansion of a CAG repeat in the first exon of the HTT gene. Affected individuals inherit more than 40 repeats and the CAG repeat is genetically unstable in both the germline and soma. Molecular diagnosis and genotyping of the CAG repeat is traditionally performed by estimation of PCR fragment size. However, this approach is complicated by the presence of an adjacent polymorphic CCG repeat and provides no information on the presence of variant repeats, flanking sequence variants or on the degree of somatic mosaicism. To overcome these limitations, we have developed an amplicon-sequencing protocol that allows the sequencing of hundreds of samples in a single MiSeq run. The composition of the HTT exon one trinucleotide repeat locus can be determined from the MiSeq sequencing reads generated. With sufficient sequencing depth, such MiSeq data can also be used to quantify the degree of somatic mosaicism of the HTT CAG repeat in the tissue analysed.

• Item Type: Research Reports or Papers (Other)
• Status: Published
• Glasgow Author(s) Enlighten ID: Hamilton, Dr Graham and Cumming, Dr Sarah and Galbraith, Ms Julie and Monckton, Professor Darren and Alshammari, Asma Mubarak and Herzyk, Dr Pawel and Mcguinness, Dr David and Ciosi, Dr Marc
• Authors: Ciosi, M., Cumming, S. A., Alshammari, A. M., Symeonidi, E., Herzyk, P., Mcguinness, D., Galbraith, J., Hamilton, G., and Monckton, D. G.
• College/School: College of Medical Veterinary and Life Sciences; College of Medical Veterinary and Life Sciences > School of Molecular Biosciences
• Publisher: Springer Nature
• Copyright Holders: Copyright © 2018 The Authors
• First Published: First posted on Protocol Exchange 2018
• Publisher Policy: Reproduced under a Creative Commons licence