Cargando…

Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing

Human pluripotent stem cells (hPSCs) have significant levels of low-grade genetic mosaicism, which commonly used techniques fail to detect in bulk DNA. These copy number variations remain a hurdle for the clinical translation of hPSC, as their effect in vivo ranges from unknown to dangerous, and the...

Descripción completa

Detalles Bibliográficos
Autores principales: Keller, Alexander, Tilleman, Laurentijn, Dziedzicka, Dominika, Zambelli, Filippo, Sermon, Karen, Van Nieuwerburgh, Filip, Spits, Claudia, Geens, Mieke
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Nature Publishing Group UK 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6796059/
https://www.ncbi.nlm.nih.gov/pubmed/31619727
http://dx.doi.org/10.1038/s41598-019-51314-6
_version_ 1783459534677737472
author Keller, Alexander
Tilleman, Laurentijn
Dziedzicka, Dominika
Zambelli, Filippo
Sermon, Karen
Van Nieuwerburgh, Filip
Spits, Claudia
Geens, Mieke
author_facet Keller, Alexander
Tilleman, Laurentijn
Dziedzicka, Dominika
Zambelli, Filippo
Sermon, Karen
Van Nieuwerburgh, Filip
Spits, Claudia
Geens, Mieke
author_sort Keller, Alexander
collection PubMed
description Human pluripotent stem cells (hPSCs) have significant levels of low-grade genetic mosaicism, which commonly used techniques fail to detect in bulk DNA. These copy number variations remain a hurdle for the clinical translation of hPSC, as their effect in vivo ranges from unknown to dangerous, and the ability to detect them will be necessary as the field advances. As such there is need for techniques which can efficiently analyse genetic content in single cells with higher throughput and lower costs. We report here on the use of the Fluidigm C1 single cell WGA platform in combination with shallow whole genome sequencing to analyse the genetic content of single hPSCs. From a hPSC line carrying an isochromosome 20, 56 single cells were analysed and found to carry a total of 50 aberrations, across 23% of cells, which could not be detected by bulk analysis. Aberrations were predominantly segmental gains, with a fewer number of segmental losses and aneuploidies. Interestingly, 40% of the breakpoints seen here correspond to known DNA fragile sites. Our results therefore demonstrate the feasibility of single cell shallow sequencing of hPSC and further expand upon the biological importance and frequency of single cell mosaicism in hPSC.
format Online
Article
Text
id pubmed-6796059
institution National Center for Biotechnology Information
language English
publishDate 2019
publisher Nature Publishing Group UK
record_format MEDLINE/PubMed
spelling pubmed-67960592019-10-25 Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing Keller, Alexander Tilleman, Laurentijn Dziedzicka, Dominika Zambelli, Filippo Sermon, Karen Van Nieuwerburgh, Filip Spits, Claudia Geens, Mieke Sci Rep Article Human pluripotent stem cells (hPSCs) have significant levels of low-grade genetic mosaicism, which commonly used techniques fail to detect in bulk DNA. These copy number variations remain a hurdle for the clinical translation of hPSC, as their effect in vivo ranges from unknown to dangerous, and the ability to detect them will be necessary as the field advances. As such there is need for techniques which can efficiently analyse genetic content in single cells with higher throughput and lower costs. We report here on the use of the Fluidigm C1 single cell WGA platform in combination with shallow whole genome sequencing to analyse the genetic content of single hPSCs. From a hPSC line carrying an isochromosome 20, 56 single cells were analysed and found to carry a total of 50 aberrations, across 23% of cells, which could not be detected by bulk analysis. Aberrations were predominantly segmental gains, with a fewer number of segmental losses and aneuploidies. Interestingly, 40% of the breakpoints seen here correspond to known DNA fragile sites. Our results therefore demonstrate the feasibility of single cell shallow sequencing of hPSC and further expand upon the biological importance and frequency of single cell mosaicism in hPSC. Nature Publishing Group UK 2019-10-16 /pmc/articles/PMC6796059/ /pubmed/31619727 http://dx.doi.org/10.1038/s41598-019-51314-6 Text en © The Author(s) 2019 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Article
Keller, Alexander
Tilleman, Laurentijn
Dziedzicka, Dominika
Zambelli, Filippo
Sermon, Karen
Van Nieuwerburgh, Filip
Spits, Claudia
Geens, Mieke
Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
title Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
title_full Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
title_fullStr Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
title_full_unstemmed Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
title_short Uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
title_sort uncovering low-level mosaicism in human embryonic stem cells using high throughput single cell shallow sequencing
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6796059/
https://www.ncbi.nlm.nih.gov/pubmed/31619727
http://dx.doi.org/10.1038/s41598-019-51314-6
work_keys_str_mv AT kelleralexander uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT tillemanlaurentijn uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT dziedzickadominika uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT zambellifilippo uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT sermonkaren uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT vannieuwerburghfilip uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT spitsclaudia uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing
AT geensmieke uncoveringlowlevelmosaicisminhumanembryonicstemcellsusinghighthroughputsinglecellshallowsequencing