Bacterial Microcolonies in Gel Beads for High-Throughput Screening of Libraries in Synthetic Biology.
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Download: Supporting_Information.pdf (469.66 [Ko])
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Serval ID
serval:BIB_6AE8F65F20EA
Type
Article: article from journal or magazin.
Collection
Publications
Institution
Title
Bacterial Microcolonies in Gel Beads for High-Throughput Screening of Libraries in Synthetic Biology.
Journal
ACS synthetic biology
ISSN
2161-5063 (Electronic)
ISSN-L
2161-5063
Publication state
Published
Issued date
17/11/2017
Peer-reviewed
Oui
Volume
6
Number
11
Pages
1988-1995
Language
english
Notes
Publication types: Journal Article
Publication Status: ppublish
Publication Status: ppublish
Abstract
Synthetic biologists increasingly rely on directed evolution to optimize engineered biological systems. Applying an appropriate screening or selection method for identifying the potentially rare library members with the desired properties is a crucial step for success in these experiments. Special challenges include substantial cell-to-cell variability and the requirement to check multiple states (e.g., being ON or OFF depending on the input). Here, we present a high-throughput screening method that addresses these challenges. First, we encapsulate single bacteria into microfluidic agarose gel beads. After incubation, they harbor monoclonal bacterial microcolonies (e.g., expressing a synthetic construct) and can be sorted according their fluorescence by fluorescence activated cell sorting (FACS). We determine enrichment rates and demonstrate that we can measure the average fluorescent signals of microcolonies containing phenotypically heterogeneous cells, obviating the problem of cell-to-cell variability. Finally, we apply this method to sort a pBAD promoter library at ON and OFF states.
Keywords
cell-to-cell variability, combinatorial libraries, directed evolution, hydrogel beads, screening, synthetic biology
Pubmed
Web of science
Open Access
Yes
Create date
09/11/2017 17:18
Last modification date
20/08/2019 14:25