@article{f804f674e55a42d1b87c5f7f1bb06f92,
title = "Pulse-Field capillary electrophoresis of repeat-primed PCR amplicons for analysis of large repeats in Spinocerebellar Ataxia Type 10",
abstract = "Large expansions of microsatellite DNA cause several neurological diseases. In Spinocerebellar ataxia type 10 (SCA10), the repeat interruptions change disease phenotype; an (ATTCC)n or a (ATCCT)n/(ATCCC)n interruption within the (ATTCT)n repeat is associated with the robust phenotype of ataxia and epilepsy while mostly pure (ATTCT)n may have reduced penetrance. Large repeat expansions of SCA10, and many other microsatellite expansions, can exceed 10,000 base pairs (bp) in size. Conventional next generation sequencing (NGS) technologies are ineffective in determining internal sequence contents or size of these expanded repeats. Using repeat primed PCR (RP-PCR) in conjunction with a high-sensitivity pulsed-field capillary electrophoresis fragment analyzer (FEMTO-Pulse, Agilent, Santa Clara, CA) (RP-FEMTO hereafter), we successfully determined sequence content of large expansion repeats in genomic DNA of SCA10 patients and transformed yeast artificial chromosomes containing SCA10 repeats. This RP-FEMTO is a simple and economical methodology which could complement emerging NGS for very long sequence reads such as Single Molecule, Real-Time (SMRT) and nanopore sequencing technologies.",
keywords = "Adult, Aged, Aged, 80 and over, Ataxin-10/genetics, DNA Repeat Expansion/genetics, Electrophoresis, Capillary/methods, Female, Humans, Male, Microsatellite Repeats/genetics, Middle Aged, Phenotype, Spinocerebellar Ataxias/genetics",
author = "Vera Hashem and Anjana Tiwari and Brittani Bewick and Teive, {Helio A.G.} and Mariana Moscovich and Birgitt Sch{\"u}ele and Khalaf Bushara and Matt Bower and Astrid Rasmussen and Tsai, {Yu Chih} and Tyson Clark and Karen McFarland and Tetsuo Ashizawa",
note = "Funding Information: Drs. Tyson Clark and Yu-Chih Tsai worked on this project without financial compensation from the NIH grant that supported this work (NS083564 to Tetsuo Ashizawa). Their contributions to this project were performed as a part of the Pacific Bioscience{\textquoteright}s own project for developing the No-Amp SMRT sequencing protocol. The collaboration was based on mutual benefits between academic investigators and PacBio investigators. PacBio provided support in the form of salaries for authors [TC and YCT], but did not have any additional role in the study design, decision to publish, or preparation of the manuscript. However, they performed No-Amp SMRT sequencing of the DNA samples that we provided and performed computational analyses of the data to generate circular consensus sequences of the repeat. The specific roles of these authors are articulated in the {\textquoteleft}author contributions{\textquoteright} section. Publisher Copyright: {\textcopyright} 2020 Hashem et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.",
year = "2020",
doi = "10.1371/journal.pone.0228789",
language = "English (US)",
volume = "15",
pages = "e0228789",
journal = "PLoS ONE",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "3",
}