On a Robust, Sensitive Cell-Free Method for Pseudomonas Sensing and Quantification in Microfluidic Templated Hydrogels
Abstract
Through the use of droplet microfluidics to integrate cell-free activity into inert hydrogel beads, we have developed a platform that can perform biologically relevant functions without the need for cells. Specifically, cell-free lysates serve a utility in performing cellular functions and providing biologically relevant metabolic products without requiring the optimal biological conditions for cell growth and proliferation. By teasing out specific biological components that enable transcription and translation to occur, these cell-like functions can be reconstituted in vitro without requiring the entire cell and milieu of cellular organelles. This enables the optimization of synthetic biological circuits, either by concentration or logic switches, simply through the addition or removal of genetic components (plasmids, inducers, or repressors) of regulatory elements. Here, we demonstrate an application of cell-free processes that is robust and portable, independent of a substrate, to apply for sensing and reporting functions of a quorum-sensing molecule N-3-oxododecanoyl homoserine lactone (3OC12HSL) found crucial for pathological Pseudomonas aeruginosa infection. We develop an agarose bead platform that is easily adaptable and simply programmable to fit a variety of biological and chemical sensing applications for the utility of ease of delivery and activation in remote environments—even in conditions with very little hydration.
- Authors:
-
[1]
- University of California at San Francisco and California, San Francisco, CA (United States). Institute for Quantitative Biosciences (QB3), Department of Bioengineering and Therapeutic Sciences; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Molecular Foundry
- Publication Date:
- Research Org.:
- Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States)
- Sponsoring Org.:
- USDOE
- OSTI Identifier:
- 1628462
- Grant/Contract Number:
- AC02-05CH11231
- Resource Type:
- Accepted Manuscript
- Journal Name:
- Micromachines
- Additional Journal Information:
- Journal Volume: 10; Journal Issue: 8; Journal ID: ISSN 2072-666X
- Publisher:
- MDPI
- Country of Publication:
- United States
- Language:
- English
- Subject:
- Science & Technology - Other Topics; Instruments & Instrumentation
Citation Formats
Seto, Jong. On a Robust, Sensitive Cell-Free Method for Pseudomonas Sensing and Quantification in Microfluidic Templated Hydrogels. United States: N. p., 2019.
Web. doi:10.3390/mi10080506.
Seto, Jong. On a Robust, Sensitive Cell-Free Method for Pseudomonas Sensing and Quantification in Microfluidic Templated Hydrogels. United States. https://doi.org/10.3390/mi10080506
Seto, Jong. Wed .
"On a Robust, Sensitive Cell-Free Method for Pseudomonas Sensing and Quantification in Microfluidic Templated Hydrogels". United States. https://doi.org/10.3390/mi10080506. https://www.osti.gov/servlets/purl/1628462.
@article{osti_1628462,
title = {On a Robust, Sensitive Cell-Free Method for Pseudomonas Sensing and Quantification in Microfluidic Templated Hydrogels},
author = {Seto, Jong},
abstractNote = {Through the use of droplet microfluidics to integrate cell-free activity into inert hydrogel beads, we have developed a platform that can perform biologically relevant functions without the need for cells. Specifically, cell-free lysates serve a utility in performing cellular functions and providing biologically relevant metabolic products without requiring the optimal biological conditions for cell growth and proliferation. By teasing out specific biological components that enable transcription and translation to occur, these cell-like functions can be reconstituted in vitro without requiring the entire cell and milieu of cellular organelles. This enables the optimization of synthetic biological circuits, either by concentration or logic switches, simply through the addition or removal of genetic components (plasmids, inducers, or repressors) of regulatory elements. Here, we demonstrate an application of cell-free processes that is robust and portable, independent of a substrate, to apply for sensing and reporting functions of a quorum-sensing molecule N-3-oxododecanoyl homoserine lactone (3OC12HSL) found crucial for pathological Pseudomonas aeruginosa infection. We develop an agarose bead platform that is easily adaptable and simply programmable to fit a variety of biological and chemical sensing applications for the utility of ease of delivery and activation in remote environments—even in conditions with very little hydration.},
doi = {10.3390/mi10080506},
journal = {Micromachines},
number = 8,
volume = 10,
place = {United States},
year = {Wed Jul 31 00:00:00 EDT 2019},
month = {Wed Jul 31 00:00:00 EDT 2019}
}
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Figure 1: Properties of the LasR-based cell-free method in bulk and in agarose bead forms (A). A schematic of the interactions between homoserine lactones and LasR proteins in relation to LasR regulation of transcription (B). A green fluorescence protein (GFP) reporter system demonstrates the bulk regulation by LasR in transcriptionalmore »
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