A Versatile Microfluidic Device for Automating Synthetic Biology
Abstract
New microbes are being engineered that contain the genetic circuitry, metabolic pathways, and other cellular functions required for a wide range of applications such as producing biofuels, biobased chemicals, and pharmaceuticals. Although currently available tools are useful in improving the synthetic biology process, further improvements in physical automation would help to lower the barrier of entry into this field. We present an innovative microfluidic platform for assembling DNA fragments with 10× lower volumes (compared to that of current microfluidic platforms) and with integrated region-specific temperature control and on-chip transformation. Integration of these steps minimizes the loss of reagents and products compared to that with conventional methods, which require multiple pipetting steps. For assembling DNA fragments, we implemented three commonly used DNA assembly protocols on our microfluidic device: Golden Gate assembly, Gibson assembly, and yeast assembly (i.e., TAR cloning, DNA Assembler). Here, we demonstrate the utility of these methods by assembling two combinatorial libraries of 16 plasmids each. Each DNA plasmid is transformed into Escherichia coli or Saccharomyces cerevisiae using on-chip electroporation and further sequenced to verify the assembly. Lastly, we anticipate that this platform will enable new research that can integrate this automated microfluidic platform to generate large combinatorial librariesmore »
- Authors:
-
- Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States). Technology Division; Sandia National Lab. (SNL-CA), Livermore, CA (United States)
- Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States). Technology Division; Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States). Fuels Synthesis Division; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division
- Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States). Fuels Synthesis Division; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division
- Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States). Fuels Synthesis Division; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division; Univ. of California, Berkeley, CA (United States). Dept. of Chemical & Biomolecular Engineering, Dept. of Bioengineering
- Joint BioEnergy Inst. (JBEI), Emeryville, CA (United States). Technology Division; Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Physical Bioscience Division
- Publication Date:
- Research Org.:
- Lawrence Berkeley National Laboratory (LBNL), Berkeley, CA (United States)
- Sponsoring Org.:
- USDOE Office of Science (SC), Biological and Environmental Research (BER)
- OSTI Identifier:
- 1378593
- Grant/Contract Number:
- AC02-05CH11231
- Resource Type:
- Accepted Manuscript
- Journal Name:
- ACS Synthetic Biology
- Additional Journal Information:
- Journal Volume: 4; Journal Issue: 10; Journal ID: ISSN 2161-5063
- Publisher:
- American Chemical Society (ACS)
- Country of Publication:
- United States
- Language:
- English
- Subject:
- 59 BASIC BIOLOGICAL SCIENCES; digital microfluidics; DNA assembly; droplet microfluidics; Gibson assembly; Golden Gate assembly; synthetic biology; TAR cloning; yeast assembly
Citation Formats
Shih, Steve C. C., Goyal, Garima, Kim, Peter W., Koutsoubelis, Nicolas, Keasling, Jay D., Adams, Paul D., Hillson, Nathan J., and Singh, Anup K. A Versatile Microfluidic Device for Automating Synthetic Biology. United States: N. p., 2015.
Web. doi:10.1021/acssynbio.5b00062.
Shih, Steve C. C., Goyal, Garima, Kim, Peter W., Koutsoubelis, Nicolas, Keasling, Jay D., Adams, Paul D., Hillson, Nathan J., & Singh, Anup K. A Versatile Microfluidic Device for Automating Synthetic Biology. United States. https://doi.org/10.1021/acssynbio.5b00062
Shih, Steve C. C., Goyal, Garima, Kim, Peter W., Koutsoubelis, Nicolas, Keasling, Jay D., Adams, Paul D., Hillson, Nathan J., and Singh, Anup K. Mon .
"A Versatile Microfluidic Device for Automating Synthetic Biology". United States. https://doi.org/10.1021/acssynbio.5b00062. https://www.osti.gov/servlets/purl/1378593.
@article{osti_1378593,
title = {A Versatile Microfluidic Device for Automating Synthetic Biology},
author = {Shih, Steve C. C. and Goyal, Garima and Kim, Peter W. and Koutsoubelis, Nicolas and Keasling, Jay D. and Adams, Paul D. and Hillson, Nathan J. and Singh, Anup K.},
abstractNote = {New microbes are being engineered that contain the genetic circuitry, metabolic pathways, and other cellular functions required for a wide range of applications such as producing biofuels, biobased chemicals, and pharmaceuticals. Although currently available tools are useful in improving the synthetic biology process, further improvements in physical automation would help to lower the barrier of entry into this field. We present an innovative microfluidic platform for assembling DNA fragments with 10× lower volumes (compared to that of current microfluidic platforms) and with integrated region-specific temperature control and on-chip transformation. Integration of these steps minimizes the loss of reagents and products compared to that with conventional methods, which require multiple pipetting steps. For assembling DNA fragments, we implemented three commonly used DNA assembly protocols on our microfluidic device: Golden Gate assembly, Gibson assembly, and yeast assembly (i.e., TAR cloning, DNA Assembler). Here, we demonstrate the utility of these methods by assembling two combinatorial libraries of 16 plasmids each. Each DNA plasmid is transformed into Escherichia coli or Saccharomyces cerevisiae using on-chip electroporation and further sequenced to verify the assembly. Lastly, we anticipate that this platform will enable new research that can integrate this automated microfluidic platform to generate large combinatorial libraries of plasmids and will help to expedite the overall synthetic biology process.},
doi = {10.1021/acssynbio.5b00062},
journal = {ACS Synthetic Biology},
number = 10,
volume = 4,
place = {United States},
year = {Mon Jun 15 00:00:00 EDT 2015},
month = {Mon Jun 15 00:00:00 EDT 2015}
}
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