Team:Toronto

From 2013.igem.org

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<h6><font color="#edebcc"><p style = "text-align:center; font-size:35px;"><b>BIOFILM ENGINEERING</b></p><br/>
<h6><font color="#edebcc"><p style = "text-align:center; font-size:35px;"><b>BIOFILM ENGINEERING</b></p><br/>
<p style = "font-size:17px;"><b><u>What are biofilms?</u></b><br/>
<p style = "font-size:17px;"><b><u>What are biofilms?</u></b><br/>
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Biofilms are communities of microbes where the cells are in an aggregate on a surface, bound together by extra-membrane material made of a mixture of specialized carbohydrates and extra-membrane proteins. The special environment in the biofilm allows cells to survive harsher environments.<br/><br/>
+
Biofilms are communities of microbes where the cells aggregate onto a surface, and are bound together by secreted extramembranous materials made of a mixture of specialised carbohydrates and extramembrane proteins. The special environment in the biofilm allows cells to survive harsher environments.<br/><br/>
<b><u>What is the “biofilm response”?</u></b><br/>
<b><u>What is the “biofilm response”?</u></b><br/>
-
Bacteria cells can exist in multiple physiological states, which are caused by environmental factors. Common ones include heat shock, nutrient (carbohydrate or amino acid) starvation, metal micronutrient starvation, etc.<br/>
+
Bacterial cells can exist in multiple physiological states, switching between them as a direct consequence of environmental factors. Common ones include heat shock, nutrient (carbohydrate or amino acid) starvation, metal micronutrient starvation, etc.<br/>
-
Gene expression in each of these physiological states is mediated by the changing the sigma factor involved in transcript synthesis. Each sigma factor has its own consensus sequence; the sigma factor on the RNA polymerase recognizes and binds to promoter regions. When a sigma factor is active, its effect is to shift which fractions of the genome are preferably expressed, because of the different promoter recognition consensus sequences. The sigma factor that is involved in the stationary phase (I.E. no motility, no reproduction) of E. coli strains is the σS factor. It directs the expression of genes necessary to induce biofilm and aggregation behavior are a subset of those genes whose promoters are recognized by the σSfactor.<br/><br/>
+
Gene expression in each of these physiological states is mediated by the changing the sigma (σ) factor involved in transcript synthesis. Each σ factor has its own consensus sequence; the σ factor on the RNA polymerase recognises and binds to promoter regions. When a σ factor is active, its effect is to shift which fractions of the genome are preferably expressed, because of the different promoter recognition consensus sequences. The sigma factor that is involved in the stationary phase (i.e. no motility, no reproduction) of E. coli strains is the σ<sup>s</sup> factor. It directs the expression of genes necessary to induce biofilm and aggregation behaviour are a subset of those genes whose promoters are recognised by the σ<sup>s</sup> factor.<br/><br/>
<b><u>Modulating and Measuring the Biofilm Response</u></b><br/>
<b><u>Modulating and Measuring the Biofilm Response</u></b><br/>
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The focus of our project is to characterize the physical and chemical manifestations of the biofilm response in cells where key genes in the biofilm response (for both structural and regulatory aspects) have been either overexpressed or deleted. The numerous assays that need to be done drove us to develop a standardized battery of assays, so that the cell can be studied as an entire system, as opposed to the methods used in past literature that only measured the effects of genetic engineering on just one manifestation. </br><br/>
+
The focus of our project is to characterise the physical and chemical manifestations of the biofilm response for cells in which key genes for both structural and regulatory aspects of the biofilm response  have been either overexpressed or deleted. The numerous assays that need to be done drove us to develop a standardised battery of assays, so that the cell can be studied as an entire system, as opposed to the methods used in past literature that only measured the effects of genetic engineering on just one manifestation of the biofilm response. </br><br/>
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<b><u>Defect in the Biobrick Paradigm</u></b><br/>
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<b><u>Defect in the BioBrick Paradigm</u></b><br/>
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The Biobrick paradigm is that gene parts are necessarily additive, due to the mechanics of the Biobrick Assembly method. Consequently, apart from by using “additive” exotic methods such as RNAi to disrupt gene expression to achieve functional gene silencing, gene deletions from a genome (negative “additive” modifications) simply cannot be submitted to the Gene Parts Registry.  
+
Due to the mechanics of the BioBrick Assembly method, the BioBrick paradigm lends itself to the construction of gene parts that are necessarily additive. Consequently, apart from using “additive” exotic methods such as RNAi to disrupt gene expression to achieve functional gene silencing, gene deletions from a genome (negative “additive” modifications) simply cannot be submitted to the Gene Parts Registry.
<br/>
<br/>
The spectacular success of the metabolic engineering field eminently shows the defect in this paradigm. Successful engineering of model organisms is based on using chemical kinetics and metabolic networks, where metabolite flux is directed by both gene up-regulation and deletion. (Insert examples here...)
The spectacular success of the metabolic engineering field eminently shows the defect in this paradigm. Successful engineering of model organisms is based on using chemical kinetics and metabolic networks, where metabolite flux is directed by both gene up-regulation and deletion. (Insert examples here...)
<br/>
<br/>
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Since deletions from a cell's genome is another means to engineering the cell as a system, we have also characterized the biofilm response of a set of knockout E. coli. Unfortunately, these systems cannot be submitted to the biobrick registry because they do not conform to the Biobrick paradigm, an issue we hope that the iGEM competition can address.
+
Since deletions from a cell's genome is another means to engineering the cell as a system, we have also characterised the biofilm response of a set of knockout E. coli. Unfortunately, these systems cannot be submitted to the BioBrick registry because they do not conform to the BioBrick paradigm, an issue we hope that the iGEM competition can address.
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Revision as of 15:16, 27 September 2013

paper

BIOFILM ENGINEERING


What are biofilms?
Biofilms are communities of microbes where the cells aggregate onto a surface, and are bound together by secreted extramembranous materials made of a mixture of specialised carbohydrates and extramembrane proteins. The special environment in the biofilm allows cells to survive harsher environments.

What is the “biofilm response”?
Bacterial cells can exist in multiple physiological states, switching between them as a direct consequence of environmental factors. Common ones include heat shock, nutrient (carbohydrate or amino acid) starvation, metal micronutrient starvation, etc.
Gene expression in each of these physiological states is mediated by the changing the sigma (σ) factor involved in transcript synthesis. Each σ factor has its own consensus sequence; the σ factor on the RNA polymerase recognises and binds to promoter regions. When a σ factor is active, its effect is to shift which fractions of the genome are preferably expressed, because of the different promoter recognition consensus sequences. The sigma factor that is involved in the stationary phase (i.e. no motility, no reproduction) of E. coli strains is the σs factor. It directs the expression of genes necessary to induce biofilm and aggregation behaviour are a subset of those genes whose promoters are recognised by the σs factor.

Modulating and Measuring the Biofilm Response
The focus of our project is to characterise the physical and chemical manifestations of the biofilm response for cells in which key genes for both structural and regulatory aspects of the biofilm response have been either overexpressed or deleted. The numerous assays that need to be done drove us to develop a standardised battery of assays, so that the cell can be studied as an entire system, as opposed to the methods used in past literature that only measured the effects of genetic engineering on just one manifestation of the biofilm response.

Defect in the BioBrick Paradigm
Due to the mechanics of the BioBrick Assembly method, the BioBrick paradigm lends itself to the construction of gene parts that are necessarily additive. Consequently, apart from using “additive” exotic methods such as RNAi to disrupt gene expression to achieve functional gene silencing, gene deletions from a genome (negative “additive” modifications) simply cannot be submitted to the Gene Parts Registry.
The spectacular success of the metabolic engineering field eminently shows the defect in this paradigm. Successful engineering of model organisms is based on using chemical kinetics and metabolic networks, where metabolite flux is directed by both gene up-regulation and deletion. (Insert examples here...)
Since deletions from a cell's genome is another means to engineering the cell as a system, we have also characterised the biofilm response of a set of knockout E. coli. Unfortunately, these systems cannot be submitted to the BioBrick registry because they do not conform to the BioBrick paradigm, an issue we hope that the iGEM competition can address.