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Team:Glasgow/Results:fixation
From 2011.igem.org
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<h2>Methods </h2> | <h2>Methods </h2> | ||
<p>We found our encapsulating material in Capsular Colanic Acid (Chao & Zhang, 2011). For simplicity it was decided to use an existing <a href="http://partsregistry.org/Part:BBa_K200000">colanic acid biobrick</a>, from the Imperial College iGEM 2009 Team.</p> | <p>We found our encapsulating material in Capsular Colanic Acid (Chao & Zhang, 2011). For simplicity it was decided to use an existing <a href="http://partsregistry.org/Part:BBa_K200000">colanic acid biobrick</a>, from the Imperial College iGEM 2009 Team.</p> | ||
| - | <p>The parts of the colanic acid system that were required to induce overexpression and export were selected. Top10 cells were transformed with the biobricks and the DNA was extracted to begin building a construct. We began attaching Ribosome Binding Sites and terminators to the biobricks we required, | + | <p>This biobrick system had been constructed to form a capsule of colanic acid that is anchored to the cell membrane by a ligase> <i>E. coli</i> naturally uses colanic acid to encapsulate itself (Stout & Gottesman, 1990), as shown in the image below. </p> |
| - | <p></p> | + | <div> |
| - | <h4>References</h4> | + | <img src="http://partsregistry.org/wiki/images/thumb/a/a3/EM.jpg/650px-EM.jpg"/> |
| + | <p><font size="1" color="grey"> Picture 1: EM image of colanic acid encapsulation of <i>E.coli</i>, showing colanic acid anchored to the membrane. (Picture as shown on BBa_K2000000 registry page)</font></p> | ||
| + | </div> | ||
| + | <p>The parts of the colanic acid system that were required to induce overexpression (Gervais, Phoenix & Drapeau, 1992) and export were selected. Anchoring to the membrane was not required. Top10 cells were transformed with the biobricks and the DNA was extracted to begin building a construct. We began attaching Ribosome Binding Sites and terminators to the biobricks we required, using the AlwNI-Method as detailed in our protocols. Due to time constraints we did not finish the entire construct. </p> | ||
| + | <p></p><h4>References</h4> | ||
<p>Chao, Y. & Zhang, T., 2011. Probing Roles of Lipopolysaccharide, Type 1 Fimbria, and Colanic Acid in the Attachment of Escherichia coli Strains on Inert Surfaces.Langmuir. 2011 Sep 20;27(18):11545-53</p> | <p>Chao, Y. & Zhang, T., 2011. Probing Roles of Lipopolysaccharide, Type 1 Fimbria, and Colanic Acid in the Attachment of Escherichia coli Strains on Inert Surfaces.Langmuir. 2011 Sep 20;27(18):11545-53</p> | ||
| + | <p>Gervais FG, Phoenix P, Drapeau GR, 1992. The rcsB gene, a positive regulator of colanic acid biosynthesis in Escherichia coli, is also an activator of ftsZ expression. Journal of Bacterioogy. 1992 Jun ; 174(12): 3964-71</p> | ||
| + | <p>Stout V, Gottesman S, 1990. RcsB and RcsC: a two-component regulator of capsule synthesis in Escherichia coli. Journal of Bacteriology. 1990 Feb ; 172(2): 659-69 </p> | ||
</html> | </html> | ||
Latest revision as of 18:31, 21 September 2011
Fixation Results
Aims
-To find a suitable material for encapsulation
-To build a construct that will appropriately express this material within a biofilm
-To transform this construct into the chassis and express it
-To characterise the expression and its effects towards fixation and encapsulation of the biofilm
Methods
We found our encapsulating material in Capsular Colanic Acid (Chao & Zhang, 2011). For simplicity it was decided to use an existing colanic acid biobrick, from the Imperial College iGEM 2009 Team.
This biobrick system had been constructed to form a capsule of colanic acid that is anchored to the cell membrane by a ligase> E. coli naturally uses colanic acid to encapsulate itself (Stout & Gottesman, 1990), as shown in the image below.
Picture 1: EM image of colanic acid encapsulation of E.coli, showing colanic acid anchored to the membrane. (Picture as shown on BBa_K2000000 registry page)
The parts of the colanic acid system that were required to induce overexpression (Gervais, Phoenix & Drapeau, 1992) and export were selected. Anchoring to the membrane was not required. Top10 cells were transformed with the biobricks and the DNA was extracted to begin building a construct. We began attaching Ribosome Binding Sites and terminators to the biobricks we required, using the AlwNI-Method as detailed in our protocols. Due to time constraints we did not finish the entire construct.
References
Chao, Y. & Zhang, T., 2011. Probing Roles of Lipopolysaccharide, Type 1 Fimbria, and Colanic Acid in the Attachment of Escherichia coli Strains on Inert Surfaces.Langmuir. 2011 Sep 20;27(18):11545-53
Gervais FG, Phoenix P, Drapeau GR, 1992. The rcsB gene, a positive regulator of colanic acid biosynthesis in Escherichia coli, is also an activator of ftsZ expression. Journal of Bacterioogy. 1992 Jun ; 174(12): 3964-71
Stout V, Gottesman S, 1990. RcsB and RcsC: a two-component regulator of capsule synthesis in Escherichia coli. Journal of Bacteriology. 1990 Feb ; 172(2): 659-69
