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Virtual Microfluidics for digital quantification and single-cell sequencing

Interest in highly parallelized analysis of single molecules and single cells is growing rapidly. Here we develop hydrogel-based virtual microfluidics as a simple alternative to complex engineered microfluidic systems for the compartmentalization of nucleic acid amplification reactions. We applied d...

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Detalles Bibliográficos
Autores principales: Xu, Liyi, Brito, Ilana L., Alm, Eric J., Blainey, Paul C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5007149/
https://www.ncbi.nlm.nih.gov/pubmed/27479330
http://dx.doi.org/10.1038/nmeth.3955
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author Xu, Liyi
Brito, Ilana L.
Alm, Eric J.
Blainey, Paul C.
author_facet Xu, Liyi
Brito, Ilana L.
Alm, Eric J.
Blainey, Paul C.
author_sort Xu, Liyi
collection PubMed
description Interest in highly parallelized analysis of single molecules and single cells is growing rapidly. Here we develop hydrogel-based virtual microfluidics as a simple alternative to complex engineered microfluidic systems for the compartmentalization of nucleic acid amplification reactions. We applied digital multiple displacement amplification (dMDA) to purified DNA templates, cultured bacterial cells, and human microbiome samples in the virtual microfluidics system and demonstrated recovery and whole-genome sequencing of single-cell MDA products. Our results from control samples showed excellent coverage uniformity and markedly reduced chimerism compared with single-cell data obtained from conventional liquid MDA reactions. We also demonstrate the applicability of the hydrogel method for genomic studies of naturally occurring microbes in human microbiome samples. The virtual microfluidics approach is a simple and robust method that will enable many laboratories to perform single-cell genomic analyses.
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spelling pubmed-50071492017-02-01 Virtual Microfluidics for digital quantification and single-cell sequencing Xu, Liyi Brito, Ilana L. Alm, Eric J. Blainey, Paul C. Nat Methods Article Interest in highly parallelized analysis of single molecules and single cells is growing rapidly. Here we develop hydrogel-based virtual microfluidics as a simple alternative to complex engineered microfluidic systems for the compartmentalization of nucleic acid amplification reactions. We applied digital multiple displacement amplification (dMDA) to purified DNA templates, cultured bacterial cells, and human microbiome samples in the virtual microfluidics system and demonstrated recovery and whole-genome sequencing of single-cell MDA products. Our results from control samples showed excellent coverage uniformity and markedly reduced chimerism compared with single-cell data obtained from conventional liquid MDA reactions. We also demonstrate the applicability of the hydrogel method for genomic studies of naturally occurring microbes in human microbiome samples. The virtual microfluidics approach is a simple and robust method that will enable many laboratories to perform single-cell genomic analyses. 2016-08-01 2016-09 /pmc/articles/PMC5007149/ /pubmed/27479330 http://dx.doi.org/10.1038/nmeth.3955 Text en Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#terms
spellingShingle Article
Xu, Liyi
Brito, Ilana L.
Alm, Eric J.
Blainey, Paul C.
Virtual Microfluidics for digital quantification and single-cell sequencing
title Virtual Microfluidics for digital quantification and single-cell sequencing
title_full Virtual Microfluidics for digital quantification and single-cell sequencing
title_fullStr Virtual Microfluidics for digital quantification and single-cell sequencing
title_full_unstemmed Virtual Microfluidics for digital quantification and single-cell sequencing
title_short Virtual Microfluidics for digital quantification and single-cell sequencing
title_sort virtual microfluidics for digital quantification and single-cell sequencing
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5007149/
https://www.ncbi.nlm.nih.gov/pubmed/27479330
http://dx.doi.org/10.1038/nmeth.3955
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