Miniaturisation of high-throughput plasmid DNA library preparation for next-generation sequencing using multifactorial optimisation

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Author(s)

Suckling, Lorna

McFarlane, Ciaran

Sawyer, Chelsea

Chambers, Stephen P

Kitney, Richard I

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Type

Journal Article

Abstract

High-throughput preparation of plasmid DNA libraries for next-generation sequencing (NGS) is an important capability for molecular biology laboratories. In particular, it is an essential quality control (QC) check when large numbers of plasmid variants are being generated. Here, we describe the use of the Design of Experiments (DOE) methodology to optimise the miniaturised preparation of plasmid DNA libraries for NGS, using the Illumina® Nextera XT technology and the Labcyte Echo® acoustic liquid dispensing system. Furthermore, we describe methods which can be implemented as a QC check for identifying the presence of genomic DNA (gDNA) in plasmid DNA samples and the subsequent shearing of the gDNA, which otherwise prevents the acoustic transfer of plasmid DNA. This workflow enables the preparation of plasmid DNA libraries which yield high-quality sequencing data.

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Date Issued

2019-03-01

Date Acceptance

2019-01-16

Citation

Synthetic and Systems Biotechnology, 2019, 4 (1), pp.57-66

URI

http://hdl.handle.net/10044/1/69621

DOI

https://www.dx.doi.org/10.1016/j.synbio.2019.01.002

ISSN

2405-805X

Publisher

KeAi

Start Page

57

End Page

66

Journal / Book Title

Synthetic and Systems Biotechnology

Volume

4

Issue

1

Copyright Statement

© 2019 Production and hosting by Elsevier B.V. on behalf of KeAi Communications Co., Ltd. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/BY-NC-ND/4.0/).

Sponsor

Engineering & Physical Science Research Council (EPSRC)

Engineering & Physical Science Research Council (EPSRC)

Biotechnology and Biological Sciences Research Council (BBSRC)

Engineering & Physical Science Research Council (EPSRC)

Identifier

http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000458661200008&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=1ba7043ffcc86c417c072aa74d649202

Grant Number

EP/L011573/1

EP/M006700/1

BB/M025632/1

EP/N023854/1

Subjects

Science & Technology

Life Sciences & Biomedicine

Biotechnology & Applied Microbiology

Publication Status

Published

Date Publish Online

2019-01-22