Difference between revisions of "Team:Yale/Design"

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        <p class="hero__desc">Developing a Framework for the Genetic Manipulation of Non-Model and Environmentally Significant Microbes</p>
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        <h1 class="hero__head">Yale iGEM</h1>
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          <h2 class="hero__year">2015</h2><a class="custom__button pdf__button" href="https://static.igem.org/mediawiki/2015/f/fb/Yale_iGEM_Project_Summary_2015.pdf">PDF Summary</a>
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            <h2 class="section__head">Project Abstract</h2>
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            <p class="abstract__text">We focused on establishing a framework to implement genetic manipulation techniques—specifically, multiplex automated genome engineering (MAGE) and CRISPR-Cas9 systems—into non-model, environmentally significant microbes using standard biological parts. The framework involves two components: (1) propagation and selection of cultures and (2) manipulation of cell genomes by MAGE and/or CRISPR. We identified design considerations for both components of the framework, and experimentally validated propagation and selection considerations using cyanobacterial strain Synechococcus sp. PCC 7002 (a fast-growing cyanobacterium capable of lipid biofuel production) and Rhizobium tropici CIAT (a nitrogen-fixing rhizobium which forms root nodules in legume plants). We then developed a workflow for the design, construction, and testing of MAGE and CRISPR technologies into non-model prokaryotes. The insights we gained from validating the propagation component of our workflow will serve to improve the versatility and robustness of our framework and will inform the development of tools for genetic manipulation in other non-model organisms.</p>
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          <h2><a href="https://2015.igem.org/Team:Yale/project" alt="Project">Project</a></h2>
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          <h2><a href="https://2015.igem.org/Team:Yale/notebook" alt="Notebook">Notebook</a></h2>
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          <h2><a href="https://2015.igem.org/Team:Yale/collaborations" alt="Collaborations">Collaborations</a></h2>
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          <h2><a href="https://2015.igem.org/Team:Yale/team" alt="Team">Team</a></h2>
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          <h2><a href="https://2015.igem.org/Team:Yale/practices" alt="Human Practices">Human Practices</a></h2>
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          <h2><a href="https://2015.igem.org/Team:Yale/standards" alt="iGEM Standards">iGEM Standards</a></h2>
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          <h2><a href="https://static.igem.org/mediawiki/2015/f/fb/Yale_iGEM_Project_Summary_2015.pdf" alt="PDF Summary">PDF Summary</a></h2>
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<p>In order to be considered for the <a href="https://2015.igem.org/Judging/Awards#SpecialPrizes">Best Applied Design award</a> and/or the <a href="https://2015.igem.org/Judging/Awards#Medals">functional prototype gold medal criterion</a>, you must fill out this page.</p>
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<p>Our project focused on developing methods to tame non-model organisms and develop them for genetic manipulation. We chose for this project Cyanobacterium and Rhizobium because we foresee their successful applications in the fields of biotechnology. Cyanobacteria can become a producer of biofuel driven by light energy and Rhizobium can cultivate the soil and control the spread of genetically modified crops.</p>
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<h5>Grow</h5>
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<p>We succeeded in growing the strains and developing a robust assay for testing growth of non-model organisms in various media. We were informed by literature and work previously done in growing these particular microbes.</p>
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<h5>Transform</h5>
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<p>We were able to transform our non-model organisms and express our heterogenous DNA. Transformation is an integral part of synthetic biology and so we spent significant effort trying multiple protocols until we arrived at one with acceptable efficiency</p>
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<h5>Select</5>
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<p>After transformation, selection was a key step. By modifying existing protocols testing for antibiotic resistance, we were able to develop a robust assay for antibiotic resistance and susceptibility in non-model organisms. This was a key step in pushing forward in taming non-model organisms.</p>
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<h5>MAGE</h5>
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<p>Multiplex automated genome engineering (MAGE) and recombineering are genetic manipulations techniques now used in E. coli, but that would make ideal techniques for use in our non-model organisms. In order to establish these in our organisms we searched through the body of research currently in existence and pulled out potentially useful recombinase proteins. We developed an assay to test for the activity of our recombinases as well.</p>
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<h5>Taming the Model Organism</h5>
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<p>Establishing genetic manipulation techniques and standard practice for these non-model organisms would have been the absolute success for our project. Thus far we have determined how to grow, transform, select, and integrate important elements of genetic manipulation systems into our organisms. More importantly, we have developed assays and principles that can be applied to the taming of any non-model organism.</p>
 
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<p>This is a prize for the team that has developed a synthetic biology product to solve a real world problem in the most elegant way. The students will have considered how well the product addresses the problem versus other potential solutions, how the product integrates or disrupts other products and processes, and how its lifecycle can more broadly impact our lives and environments in positive and negative ways.</p>
 
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If you are working on art and design as your main project, please join the art and design track. If you are integrating art and design into the core of your main project, please apply for the award by completing this page.
 
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Revision as of 03:47, 19 September 2015


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Developing a Framework for the Genetic Manipulation of Non-Model and Environmentally Significant Microbes

Yale iGEM

2015

PDF Summary

Project Abstract

We focused on establishing a framework to implement genetic manipulation techniques—specifically, multiplex automated genome engineering (MAGE) and CRISPR-Cas9 systems—into non-model, environmentally significant microbes using standard biological parts. The framework involves two components: (1) propagation and selection of cultures and (2) manipulation of cell genomes by MAGE and/or CRISPR. We identified design considerations for both components of the framework, and experimentally validated propagation and selection considerations using cyanobacterial strain Synechococcus sp. PCC 7002 (a fast-growing cyanobacterium capable of lipid biofuel production) and Rhizobium tropici CIAT (a nitrogen-fixing rhizobium which forms root nodules in legume plants). We then developed a workflow for the design, construction, and testing of MAGE and CRISPR technologies into non-model prokaryotes. The insights we gained from validating the propagation component of our workflow will serve to improve the versatility and robustness of our framework and will inform the development of tools for genetic manipulation in other non-model organisms.

Find Out More

Project

Notebook

Collaborations

Team

Human Practices

iGEM Standards

PDF Summary

Design

By talking about your design work on this page, there is one medal criterion that you can attempt to meet, and one award that you can apply for. If your team is going for a gold medal by building a functional prototype, you should tell us what you did on this page. If you are going for the Applied Design award, you should also complete this page and tell us what you did.

Project Design

Our project focused on developing methods to tame non-model organisms and develop them for genetic manipulation. We chose for this project Cyanobacterium and Rhizobium because we foresee their successful applications in the fields of biotechnology. Cyanobacteria can become a producer of biofuel driven by light energy and Rhizobium can cultivate the soil and control the spread of genetically modified crops.

Grow

We succeeded in growing the strains and developing a robust assay for testing growth of non-model organisms in various media. We were informed by literature and work previously done in growing these particular microbes.

Transform

We were able to transform our non-model organisms and express our heterogenous DNA. Transformation is an integral part of synthetic biology and so we spent significant effort trying multiple protocols until we arrived at one with acceptable efficiency

Select

After transformation, selection was a key step. By modifying existing protocols testing for antibiotic resistance, we were able to develop a robust assay for antibiotic resistance and susceptibility in non-model organisms. This was a key step in pushing forward in taming non-model organisms.

MAGE

Multiplex automated genome engineering (MAGE) and recombineering are genetic manipulations techniques now used in E. coli, but that would make ideal techniques for use in our non-model organisms. In order to establish these in our organisms we searched through the body of research currently in existence and pulled out potentially useful recombinase proteins. We developed an assay to test for the activity of our recombinases as well.

Taming the Model Organism

Establishing genetic manipulation techniques and standard practice for these non-model organisms would have been the absolute success for our project. Thus far we have determined how to grow, transform, select, and integrate important elements of genetic manipulation systems into our organisms. More importantly, we have developed assays and principles that can be applied to the taming of any non-model organism.