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The Development of an Effective Bacterial Single-Cell Lysis Method Suitable for Whole Genome Amplification in Microfluidic Platforms

Liu, Y. and Schulze-Makuch, D. and de Vera, J. P. and Cockell, C. and Leya, T. and Baque, Mickael and Walther-Antonio, M. (2018) The Development of an Effective Bacterial Single-Cell Lysis Method Suitable for Whole Genome Amplification in Microfluidic Platforms. Micromachines, pp. 1-18. Multidisciplinary Digital Publishing Institute (MDPI). DOI: 10.3390/mi9080367 ISSN 2072-666X

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Official URL: http://www.mdpi.com/2072-666X/9/8/367

Abstract

Single-cell sequencing is a powerful technology that provides the capability of analyzing a single cell within a population. This technology is mostly coupled with microfluidic systems for controlled cell manipulation and precise fluid handling to shed light on the genomes of a wide range of cells. So far, single-cell sequencing has been focused mostly on human cells due to the ease of lysing the cells for genome amplification. The major challenges that bacterial species pose to Genome amplification from single cells include the rigid bacterial cell walls and the need for an effective Lysis protocol compatible with microfluidic platforms. In this work, we present a lysis protocol that can be used to extract genomic DNA from both gram-positive and gram-negative species without interfering with the amplification chemistry. Corynebacterium glutamicum was chosen as a typical gram-positive model and Nostoc sp. as a gram-negative model due to major challenges reported in previous studies. Our protocol is based on thermal and chemical lysis. We consider 80% of single-cell replicates that lead to >5 ng DNA after amplification as successful attempts. The protocol was directly applied to Gloeocapsa sp. and the single cells of the eukaryotic Sphaerocystis sp. and achieved a 100% success rate.

Item URL in elib:https://elib.dlr.de/121431/
Document Type:Article
Title:The Development of an Effective Bacterial Single-Cell Lysis Method Suitable for Whole Genome Amplification in Microfluidic Platforms
Authors:
AuthorsInstitution or Email of AuthorsAuthors ORCID iD
Liu, Y.Department of Surgery, Division of Surgical Research, Mayo Clinic, Rochester, MN 55905, USAUNSPECIFIED
Schulze-Makuch, D.TU Berlin, GermanyUNSPECIFIED
de Vera, J. P.DLR, German Aerospace Center, Management and Infrastructure, Astrobiological Laboratories, Berlin, Germany.https://orcid.org/0000-0002-9530-5821
Cockell, C.UK Centre for Astrobiology, School of Physics and Astronomy, University of Edinburgh, Edinburgh, UK; e-mail: c.s.cockell (at) ed.ac.ukUNSPECIFIED
Leya, T.Fraunhofer IZI-BB, Potsdam, GermanyUNSPECIFIED
Baque, MickaelMickael.Baque (at) dlr.dehttps://orcid.org/0000-0002-6696-6030
Walther-Antonio, M.UNSPECIFIEDUNSPECIFIED
Date:25 July 2018
Journal or Publication Title:Micromachines
Refereed publication:Yes
Open Access:Yes
Gold Open Access:Yes
In SCOPUS:Yes
In ISI Web of Science:Yes
DOI :10.3390/mi9080367
Page Range:pp. 1-18
Publisher:Multidisciplinary Digital Publishing Institute (MDPI)
ISSN:2072-666X
Status:Published
Keywords:bacteria lysis protocol; microalgae lysis; single-cell multiple displacement amplification
HGF - Research field:Aeronautics, Space and Transport
HGF - Program:Space
HGF - Program Themes:Space Science and Exploration
DLR - Research area:Raumfahrt
DLR - Program:R EW - Erforschung des Weltraums
DLR - Research theme (Project):R - Vorhaben Planetary Evolution and Life
Location: Berlin-Adlershof
Institutes and Institutions:Institute of Planetary Research > Leitungsbereich PF
Deposited By: de Vera, Dr. Jean Pierre Paul
Deposited On:10 Sep 2018 16:13
Last Modified:20 Jan 2020 13:35

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