Pulse-Field capillary electrophoresis of repeat-primed PCR amplicons for analysis of large repeats in Spinocerebellar Ataxia Type 10.

Autor: Hashem V; Department of Neurology, Houston Methodist Research Institute, Houston, Texas, United States of America., Tiwari A; Department of Neurology, Houston Methodist Research Institute, Houston, Texas, United States of America., Bewick B; Department of Neurology, Houston Methodist Research Institute, Houston, Texas, United States of America., Teive HAG; Movement Disorders Unit, Neurology Service, Department of Internal Medicine, Hospital de Clinicas, Federal University of Paraná, Curitiba, PR, Brazil., Moscovich M; Department of Internal Medicine, Federal University of Paraná, Curitiba, Paraná, Brazil., Schüele B; Department of Pathology, Stanford University School of Medicine, Stanford, California, United States of America., Bushara K; Department of Neurology, University of Minnesota Medical Center, Minneapolis, Minnesota, United States of America., Bower M; Institute of Human Genetics, University of Minnesota Medical Center, Minneapolis, Minnesota, United States of America., Rasmussen A; Oklahoma Medical Research Foundation, Oklahoma City, OK, USA.; Instituto Nacional de Ciencias Medicas y Nutricion Salvador Zubiran, Mexico City, CDMX, Mexico., Tsai YC; Pacific Biosciences of California, Inc, Menlo Park, California, United States of America., Clark T; Pacific Biosciences of California, Inc, Menlo Park, California, United States of America., McFarland K; Department of Neurology and The McKnight Brain Institute, University of Florida, Gainesville, Florida, United States of America., Ashizawa T; Department of Neurology, Houston Methodist Research Institute, Houston, Texas, United States of America.
Jazyk: angličtina
Zdroj: PloS one [PLoS One] 2020 Mar 11; Vol. 15 (3), pp. e0228789. Date of Electronic Publication: 2020 Mar 11 (Print Publication: 2020).
DOI: 10.1371/journal.pone.0228789
Abstrakt: 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.
Competing Interests: 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. This does not alter the authors' adherence to PLOS ONE policies on sharing data and materials.
Databáze: MEDLINE
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