Team:Newcastle

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                 <h3>Our Project</h3>
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Our project focuses on the creation and applications of L-forms: bacteria that grow without a cell wall. We propose L-forms as a novel chassis for synthetic biology. Our principle BioBrick switches <i>Bacillus subtilis</i> cells between rod-shape and L-form.
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We will use microfluidics to attempt genome shuffling and shape-shifting. It is easier to fuse bacteria without cell walls. Fusion will cause genetic recombination, allowing directed evolution. We will put L-forms in moulds to observe if they adopt different shapes.  
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L-forms exist symbiotically in plants, which we will visualise by growing GFP labelled L-forms inside seedlings. L-forms could be engineered to supply nutrients to their host. L-forms are osmotically sensitive, giving biosecurity that they lyse if they escape from the plant.
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As outreach we reflected upon our project's implications with stakeholders, created a BioGame for the public and developed a workshop for those new to modelling. Finally, we evaluated the relationship between synthetic biology and architecture.
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                    Click below to explore!
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                <h3>Rod to L-form Switch BioBrick</h3>
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                <p>An L-form is a bacterium that has no cell wall but is still able to multiply. Many species including <i>Bacillus subtilis</i> and <i>Escherichia coli</i> have an L-form. Losing the cell wall alters the characteristics of a bacterium creating a wealth of new applications for a bacterial species. In order to produce L-forms we have developed a BioBrick to allow the switching between the rod and L-form state of <i>B. subtilis</i>.</p>
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                <p>This BioBrick integrates into the <i>B. subtilis</i> chromosome by homologous recombination.  It replaces the region containing the end of pbpb gene and the beginning of the <i>murE </i> gene (<i>murE </i> enables cell wall biosynthesis). However, it replaces the constitutive <i>murE </i> promoter with a xylose-inducible promoter (<i>PxylR</i>).  When xylose is present cell wall biosynthesis is switched on, giving rod cells. When xylose is not present, the cells will become L-forms.</p>
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                 <h2>Our Project</h2>
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                 <h3>Shape Shifting</h3>
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                Our project focuses on the creation and applications of L-forms: bacteria that grow without a cell wall. We propose L-forms as a novel chassis for synthetic biology. Our principle BioBrick switches <i>Bacillus subtilis</i> cells between rod-shape and L-form.
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The loss of the cell wall leaves L-forms protected by only a fluid cell membrane. This allows L-forms to adapt to shapes of cracks and cavities, and possibly squeeze through tiny channels to deliver cargo to otherwise inaccessible targets.
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                We will use microfluidics to attempt genome shuffling and shape-shifting. It is easier to fuse bacteria without cell walls. Fusion will cause genetic recombination, allowing directed evolution. We will put L-forms in moulds to observe if they adopt different shapes.
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We will inject L-forms into specially designed microfluidics chambers and observe their behaviour and shape. We are modelling membrane behaviour under stress conditions, which includes cell growth and boundaries provided by the chamber walls.  
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                L-forms exist symbiotically in plants, which we will visualise by growing GFP labelled L-forms inside seedlings. L-forms could be engineered to supply nutrients to their host. L-forms are osmotically sensitive, giving biosecurity that they lyse if they escape from the plant.
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                As outreach we reflected upon our project's implications with stakeholders, created a BioGame for the public and developed a workshop for those new to modelling. Finally, we evaluated the relationship between synthetic biology and architecture.
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                 <p>L-forms and plants can exist in a symbiotic relationship as plants provide an osmotically suitable environment. In return, L-forms can confer benefits to their host including reducing the rate of fungal infection. We plan to wash seedlings in a solution of GFP labelled L-forms, allowing the seedlings to take up the bacteria. We will then view our L-forms inside the plant using confocal microscopy.
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In the future, L-forms could be engineered to supply nutrients to plants, potentially increasing crop yield in low fertility soil. L-forms are osmotically sensitive, giving the ethical advantage that they lyse if they escape from their host plant into the environment. Please click this box for more information.
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We have created a novel chassis for Synthetic Biology, one that is not hindered by its cell wall. These cells are called “L-Forms” or as we prefer to call them: naked bacteria. Even better, we’ve already begun putting them to good use!
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Many of the interactions between engineering and biology, required in Synthetic Biology, would be made easier without the barrier of the cell wall.  Bacteria with a cell wall are harder to get things into and out of, harder to fuse together and won’t mould into different shapes. Our main BioBrick allows us not only to remove the cell wall, but to turn it back on again at the flick of a switch, ensuring the bacterium’s dignity remains intact!
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We can fuse L-forms together and recombine their genomes - our naked bacteria undergo sexual reproduction! ;) This can be used to shuffle genomes and perform directed evolution to produce bacteria with improved phenotypes.
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L-forms have been shown to inhabit plants, we didn’t want our naked bacteria to feel left out so we put them inside plants too. They could provide natural resistance to the plant from  pathogens, and could be used to deliver  useful molecules.
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Naked bacteria cannot live outside of an osmotically suitable environment; as soon as they leave a plant or the lab, they’ll burst. They essentially have an in-built kill-switch. You won’t be finding any of our naked bacteria getting dirty in soil.
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                <h3>Genome shuffling</h3>
 
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                <p>Genome shuffling is an innovative way to select for improvement in desired traits, including survival in harsh conditions and increased protein production. One method involves squashing two genetically similar cells together until they fuse. This results in the swapping of sections between each genome, producing two new genetic mosaics. These can then be assayed for desired traits, and the procedure repeated. </p>
 
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<p>L-forms do not have cells walls so are comparatively easy to fuse using microfluidics. We have created BioBricks encoding green and red fluorescently labelled DNA binding proteins. Fusing one GFP and one RFP chromosome labelled L-form will allow us to visualize the swapping of DNA between the bacteria: the new cells will fluoresce with both red and green.
 
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Latest revision as of 12:09, 26 October 2013

 
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IGEM Home Newcastle University

Click below to explore!

Our Project

We have created a novel chassis for Synthetic Biology, one that is not hindered by its cell wall. These cells are called “L-Forms” or as we prefer to call them: naked bacteria. Even better, we’ve already begun putting them to good use!

Many of the interactions between engineering and biology, required in Synthetic Biology, would be made easier without the barrier of the cell wall. Bacteria with a cell wall are harder to get things into and out of, harder to fuse together and won’t mould into different shapes. Our main BioBrick allows us not only to remove the cell wall, but to turn it back on again at the flick of a switch, ensuring the bacterium’s dignity remains intact!

We can fuse L-forms together and recombine their genomes - our naked bacteria undergo sexual reproduction! ;) This can be used to shuffle genomes and perform directed evolution to produce bacteria with improved phenotypes.

L-forms have been shown to inhabit plants, we didn’t want our naked bacteria to feel left out so we put them inside plants too. They could provide natural resistance to the plant from pathogens, and could be used to deliver useful molecules.

Naked bacteria cannot live outside of an osmotically suitable environment; as soon as they leave a plant or the lab, they’ll burst. They essentially have an in-built kill-switch. You won’t be finding any of our naked bacteria getting dirty in soil.

Newcastle University The Centre for Bacterial Cell Biology Newcastle Biomedicine The School of Computing Science The School of Computing Science

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