Isolation and Characterization of Bacterial Cellulase Producers for Biomass Deconstruction: A Microbiology Laboratory Course †
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Authors:
Jesus F. Barajas1,2,
Maren Wehrs2,3,
Milton To2,3,
Lauchlin Cruickshanks4,
Rochelle Urban2,3,5,
Adrienne McKee2,3,6,
John Gladden7,
Ee-Been Goh2,3,8,
Margaret E. Brown2,3,9,
Diane Pierotti2,3,
James M. Carothers10,
Aindrila Mukhopadhyay2,3,
Jay D. Keasling2,3,10,11,12,13,14,15,
Jeffrey L. Fortman2,3,15,
Steven W. Singer2,3,*,
Constance B. Bailey2,3,11,#,*
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Received 11 December 2018 Accepted 22 February 2019 Published 26 July 2019
- ©2019 Author(s). Published by the American Society for Microbiology
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[open-access] This is an Open Access article distributed under the terms of the Creative Commons Attribution-Noncommercial-NoDerivatives 4.0 International license (https://creativecommons.org/licenses/by-nc-nd/4.0/ and https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode), which grants the public the nonexclusive right to copy, distribute, or display the published work.
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†Supplemental materials available at http://asmscience.org/jmbe
- *Corresponding author. Mailing address: Joint BioEnergy Institute, 5885 Hollis St, 4th Floor, Emeryville, CA 94608. Phone: 510-486-5556. E-mail: [email protected].
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# Current address: University of Tennessee-Knoxville, Knoxville, 1420 Circle Drive, Knoxville, TN 37996. Phone 865-974-8378. E-mail: [email protected].
Abstract:
The conversion of biomass to biofuels presents a solution to one of the largest global challenges of our era, climate change. A critical part of this pipeline is the process of breaking down cellulosic sugars from plant matter to be used by microbes containing biosynthetic pathways that produce biofuels or bioproducts. In this inquiry-based course, students complete a research project that isolates cellulase-producing bacteria from samples collected from the environment. After obtaining isolates, the students characterize the production of cellulases. Students then amplify and sequence the 16S rRNA genes of confirmed cellulase producers and use bioinformatic methods to identify the bacterial isolates. Throughout the course, students learn about the process of generating biofuels and bioproducts through the deconstruction of cellulosic biomass to form monosaccharides from the biopolymers in plant matter. The program relies heavily on active learning and enables students to connect microbiology with issues of sustainability. In addition, it provides exposure to basic microbiology, molecular biology, and biotechnology laboratory techniques and concepts. The described activity was initially developed for the Introductory College Level Experience in Microbiology (iCLEM) program, a research-based immersive laboratory course at the US Department of Energy Joint BioEnergy Institute. Originally designed as an accelerated program for high-potential, low-income, high school students (11th–12th grade), this curriculum could also be implemented for undergraduate coursework in a research-intensive laboratory course at a two- or four-year college or university.
References & Citations
Supplemental Material
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Appendix 1: List of reagents and instrumentation, Appendix 2: Faculty instructions, Appendix 3: Student handouts and protocols, Appendix 4: Recipes and detailed protocols, Appendix 5: Glossary and additional references, Appendix 6: Worksheets and templates, Appendix 7: Recommended exam questions and rubric, Appendix 8: Module pre-/post-assessment questions and rubric
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Abstract:
The conversion of biomass to biofuels presents a solution to one of the largest global challenges of our era, climate change. A critical part of this pipeline is the process of breaking down cellulosic sugars from plant matter to be used by microbes containing biosynthetic pathways that produce biofuels or bioproducts. In this inquiry-based course, students complete a research project that isolates cellulase-producing bacteria from samples collected from the environment. After obtaining isolates, the students characterize the production of cellulases. Students then amplify and sequence the 16S rRNA genes of confirmed cellulase producers and use bioinformatic methods to identify the bacterial isolates. Throughout the course, students learn about the process of generating biofuels and bioproducts through the deconstruction of cellulosic biomass to form monosaccharides from the biopolymers in plant matter. The program relies heavily on active learning and enables students to connect microbiology with issues of sustainability. In addition, it provides exposure to basic microbiology, molecular biology, and biotechnology laboratory techniques and concepts. The described activity was initially developed for the Introductory College Level Experience in Microbiology (iCLEM) program, a research-based immersive laboratory course at the US Department of Energy Joint BioEnergy Institute. Originally designed as an accelerated program for high-potential, low-income, high school students (11th–12th grade), this curriculum could also be implemented for undergraduate coursework in a research-intensive laboratory course at a two- or four-year college or university.

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Author and Article Information
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Received 11 December 2018 Accepted 22 February 2019 Published 26 July 2019
- ©2019 Author(s). Published by the American Society for Microbiology
-
[open-access] This is an Open Access article distributed under the terms of the Creative Commons Attribution-Noncommercial-NoDerivatives 4.0 International license (https://creativecommons.org/licenses/by-nc-nd/4.0/ and https://creativecommons.org/licenses/by-nc-nd/4.0/legalcode), which grants the public the nonexclusive right to copy, distribute, or display the published work.
-
†Supplemental materials available at http://asmscience.org/jmbe
- *Corresponding author. Mailing address: Joint BioEnergy Institute, 5885 Hollis St, 4th Floor, Emeryville, CA 94608. Phone: 510-486-5556. E-mail: [email protected].
-
# Current address: University of Tennessee-Knoxville, Knoxville, 1420 Circle Drive, Knoxville, TN 37996. Phone 865-974-8378. E-mail: [email protected].
Figures

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FIGURE 1
Biomass to biofuels and bioproducts pipeline. The activity described falls under deconstruction, the process of discovering enzymes that break down polysaccharides to form sugars and lignin-derived intermediates (indicated by orange text and box) that can be metabolized by bacteria to generate petrochemical replacements.

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FIGURE 2
Student confidence assessment for learning objective 4 (see Table 1 ). Survey was on a scale of 1 to 10, with 1 being the least confident (Very Low) and 10 being the most confident (Very High).