From 2011.igem.org

Information about our novel biofilm forming chassis, E.coli Nissle 1917 is also included here. (Safety|More info)
Please click on either the safety or info tabs to find out more about each of these biobricks.

Biobrick: Phosphodiesterase (PDE)

Host:
E.coli Top 10

Origin:
Amplified from the Pseudomonas aeruginosa PAO1 genome

Function & Use:
Phosphodiesterases are a family of enzymes naturally present in microorganisms, which break phosphodiester bonds. The specific enzyme we are using breaks the phosphodiester bond in the second messenger nucleotide, cyclic digaunylate (cyclic-di-GMP). Cyclic-di-GMP is important in many bacterial processes, including biofillm formation and motility. The enzyme regulates signal transduction by controlling levels of the signalling molecule in cells.

We are using the enzyme to control the levels of cyclic-di-GMP within the cell. We expect that the targeted expression of phosphodesiterase could be used to interfere with biofilm formation or to trigger dispersal.

Safety:
Phosphodiesterase enzymes are naturally present in many organisms, including E.coli. We are using it to disrupt biofilm formation by disrupting levels of cyclic-di-GMP, which is the natural function of the enzyme and has already been studied. We do not anticipate the phosphodiesterase biobrick posing a safety risk.

References & Further Reading:

"The EAL Domain Protein VieA Is a Cyclic Diguanylate Phosphodiesterase" which can be accessed here.

"A blue light-inducible phosphodiesterase activity in the cyanobacterium Synechococcus elongates" which can be accessed here.

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Biobrick: LOV2

Host:
E.coli (Top 10)

Origin:
The LOV domain was originally isolated from Arabidopsis. It was characterised by Dr John Christie of the University of Glasgow.

The version of LOV2 we are using was codon optimised for expression in E.coli and synthesized by researchers from the University of Glasgow’s Institute Of Infection, Immunity and Inflammation.

Function & Uses:
The LOV (Light-Oxygen-Voltage) domain is a photoreceptor that responds to blue light. In nature it was first found to be involved in the phototropism response in plants and has since been found to be present in fungi and bacteria also. It has been shown to be coupled to many domains, for example phosphodiesterase or kinases.

We are using it in our cells as a reporter due to its ability to function in anoxic conditions. This is particularly useful in biofilms and is a function that fluorescent proteins derived from GFP do not have.

Safety:
It has not been attached to any functional domain and the LOV domain itself has no pathogenic properties. We therefore do not expect the LOV2 domain to have any safety risks.

It has previously been expressed in E.coli by Dr John Christie.

References & Further Reading:

"LOV (light, oxygen, or voltage) domains of the blue-light photoreceptor phototropin (nph1): Binding sites for the chromophore flavin mononucleotide" which can be accessed here.

"The LOV Domain Family: Photoresponsive Signaling Modules Coupled to Diverse Output Domains" which can be accessed here.

"Information on the LOV Domain" by Dr John Christie which can be accessed here.

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Biobrick: iLOV

Host:
E.coli (Top 10)

Origin:
iLOV is a version of LOV2 that has been altered through site directed mutagenesis and DNA shuffling.

Function & Use
iLOV has the same function and uses as LOV2. As a reporter, it is advantageous over GFP derived fluorescent proteins due to its small size (useful if you are tagging proteins), ability to recover quickly from photobleaching and use in anoxic conditions.

Safety:
The mutations that iLOV has the function of increasing intensity of fluorescence only. It does not have any altered biological role or new biological function.

Therefore we do not antipate any way in which it could increase pathogenicity or survival in the wild.
References and Further Reading:

"The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection" which can be accessed here.

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Biobrick: Ranaspumin (RSN2)

Host:
E.coli (Top 10)

Origin:
Túngara Frog (Engystomops pustulosus)

Function & Uses:
Ranaspumin is a surfactant protein found in the foam nests of the Túngara frog. In nature it is used to protect and incubate the fertilized eggs of the frog. It has natural antimicrobial functions and antibiofilm activity has been reported.

Safety:
We are using it to aid in dispersal of biofilms, which is a natural property of the protein. It has no known biological risk to humans or the environment.

References & Further Reading
"Ranaspumin-2: Structure and Function of a Surfactant Protein from the Foam Nests of a Tropical Frog" which can be accessed here.

"Biofoams and natural protein surfactants" which can be accessed here.

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Biobrick: Ranaspumin with 6xHIS tag

We have also created a version of the Ranaspumin biobrick with a 6xHIS tag.

The HIS tag would not give the microorganism any enhanced ability to survive outside the lab, or pose a safety risk to the public or environment.

References & Further Reading

Information on "His-tagged proteins", which can be accessed here.

Information on "Affinity Purification", which can be accessed here.

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Biobrick: Latherin

Host: E.coli Top 10

Origin: Horse (Equus caballas)

Function & Uses:
Latherin is a surfactant protein that was originally isolated from horse sweat. Its normal biological function is temperature regulation and is believed to function by enhancing evaporation from the pelt. Due to its ability to bind to hydrophobic surfaces, the protein is being investigated for its possible ability to aid in breaking up biofilms.

Safety:
We are using latherin to aid in dispersal of biofilms, which is a natural property of the protein that has already been investigated.

Latherin is suspected to be involved in the allergen response to horses. We propose that anything containing the protein should be clearly labelled as such, so that those with a horse allergy can avoid contact with it.

References & Further Reading:

"Latherin: A Surfactant Protein of Horse Sweat and Saliva" which can be accessed here

"Isolation and characterization of latherin, a surface-active protein from horse sweat" which can be accessed here.

"Biochemical characterization and surfactant properties of horse allergens" which can be accessed here.

"Latherin and other biocompatible surfactant proteins" which can be accessed here

Return to top

Biobrick: Latherin with 6xHIS tag

We have also created a version of the Latherin biobrick with 6xHIS tag for affinity purification.

The use of 6xHIS tags is well established and would not give the microorganism any enhanced ability to survive outside the lab, or pose a safety risk to the public or environment.

References & Further Reading

Information on "His-tagged proteins", which can be accessed here.

Information on "Affinity Purification", which can be accessed here.

Return to top

Nissle 1917 Strain of E.coli

Nissle 1917 is a strain of E.coli that we are working with due to its ability to form biofilms. This is a capacity that laboratory strains of E.coli have lost.

It is used as a probiotic in health supplements, which are available to buy commercially in Germany as Mutaflor tablets. It is also studied as a carrier for therapeutic molecules, due to the fact it is safe for human consumption and does not cause illness when it colonises the gut.

References & Further Reading:
"Intestinal immunity of Escherichia coli NISSLE 1917: a safe carrier for therapeutic molecules" which can be accessed here.

"Maintaining remission of ulcerative colitis with the probiotic Escherichia coli Nissle 1917 is as effective as with standard mesalazine" which can be accessed here.

"NF- B- and AP-1-Mediated Induction of Human Beta Defensin-2 in Intestinal Epithelial Cells by Escherichia coli Nissle 1917: a Novel Effect of a Probiotic Bacterium" which can be accessed here.

@@ Line 1: / Line 1: @@
 {{Team:Glasgow/Header}}
 <html>
-<h1>Biobrick: Phosphodiesterase (PDE)</h1>
+<a name="top"></a><h1>Biobrick Safety</h1>
+<p>Here you can find safety information on the following novel biobricks:</p>
+<ul>Phosphodiesterase (<a href="#pde">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/PDE">More info</a>)</ul>
+<ul>LOV2 (<a href="#lov2">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/LOV2">More info</a>)</ul>
+<ul>iLOV (<a href="#ilov">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/LOV2">More info</a>)</ul>
+<ul>Ranaspumin (<a href="#rana">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/Ranaspumin2">More info</a>)</ul>
+<ul>Ranaspumin with 6xHIS tag (<a href="#rana6">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/Ranaspumin2">More info</a>)</ul>
+<ul>Latherin (<a href="#lath">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/Parts/Latherin">More info</a>)</ul>
+<ul>Latherin with 6xHIS tag (<a href="#lath6">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/Parts/Latherin">More info</a>)</ul>
+<br/>
+<p>Information about our novel biofilm forming chassis, <i>E.coli</i> Nissle 1917 is also included here. (<a href="#nis">Safety</a>|<a href="https://2011.igem.org/Team:Glasgow/Nissle">More info</a>)<br/>Please click on either the safety or info tabs to find out more about each of these biobricks.</p>
+<a name="pde"></a><h2>Biobrick: Phosphodiesterase (PDE)</h2>
 <b>Host:</b><br />
@@ Line 7: / Line 18: @@
 <b>Origin:</b><br />
-<i>Pseudomonas aeruginosa</i> PAO1<br /><br />
+Amplified from the <i>Pseudomonas aeruginosa</i> PAO1 genome<br /><br />
 <b>Function & Use:</b><br />
-Phosphodiesterase is an enzyme naturally present in microorganisms, which breaks phosphodiester bonds.  The specific enzyme we are using is a cyclic nucleotide phosphodiesterase, meaning that it breaks the phosphodiesterase bond in second messenger nucleotide molecules, such as cAMP and cGMP. Its role is to regulate signal transduction by controlling levels of signalling molecules in cells.<br /> <br />
+Phosphodiesterases are a family of enzymes naturally present in microorganisms, which break phosphodiester bonds.  The specific enzyme we are using breaks the phosphodiester bond in the second messenger nucleotide, cyclic digaunylate (cyclic-di-GMP).  Cyclic-di-GMP  is important in many bacterial processes, including biofillm formation and motility.  The enzyme regulates signal transduction by controlling levels of the signalling molecule in cells. <br /> <br />
-We are specifically using it to break down the signalling molecule cyclic diguanylate (cyclic-di-GMP).  The aim of this is to use it control the levels of cyclic-di-GMP within the cell, which acts as a second messenger and is key to many bacterial processes such as biofilm formation and motility.  We therefore expect that the targeted expression of phosphodesiterase could be used to interfere with biofilm formation or to trigger dispersal.<br /><br />
+We are using the enzyme to control the levels of cyclic-di-GMP within the cell.  We expect that the targeted expression of phosphodesiterase could be used to interfere with biofilm formation or to trigger dispersal.<br /><br />
 <b>Safety:</b><br />
@@ Line 19: / Line 30: @@
 Phosphodiesterase enzymes are naturally present in many organisms, including <i>E.coli.</i>  We are using it to disrupt biofilm formation by disrupting levels of cyclic-di-GMP, which is the natural function of the enzyme and has already been studied.  We do not anticipate the phosphodiesterase biobrick posing a safety risk.<br /><br />
-<b>References & Further Reading:</b><br/>
+<b>References & Further Reading:</b><br/><br/>
-"The EAL Domain Protein VieA Is a Cyclic Diguanylate Phosphodiesterase" which can be accessed here <a href=http://www.jbc.org/content/280/39/33324.abstract>here</a><br /><br />
+"The EAL Domain Protein VieA Is a Cyclic Diguanylate Phosphodiesterase" which can be accessed <a href=http://www.jbc.org/content/280/39/33324.abstract>here.</a><br /><br />
-"A blue light-inducible phosphodiesterase activity in the cyanobacterium <i>Synechococcus elongates</i>" which can be accessed here <a href=http://www.ncbi.nlm.nih.gov/pubmed/20408974>here</a>
+"A blue light-inducible phosphodiesterase activity in the cyanobacterium <i>Synechococcus elongates</i>" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/pubmed/20408974>here.</a>
-<h1>Biobrick: Phosphodiesterase with 6xHIS Tag</h1>
-We have also created a version of the phosphodiesterase biobrick with 6xHIS tag for affinity purification.<br /><br />
-The use of 6xHIS tags is well established and would not give the microorganism any enhanced ability to survive outside the lab, or pose a safety risk to the public or environment. <br /><br />
-<b>References & Further Reading:</b><br/>
+<h6><a href="#top">Return to top</a></h6>
-"Making the most of affinity tags" which can be accessed <a href=http://www.sciencedirect.com/science/article/pii/S0167779905000843>here</a><br/>
+<a name="lov2"></a><h2>Biobrick: LOV2</h2>
-<h1>Biobrick: LOV2</h1>
 <b>Host:</b><br />
@@ Line 52: / Line 56: @@
 <b>Safety:</b><br />
-It has not been attached to any functional domain and the LOV domain itself has no pathogenic properties.  We therefore do not expect the LOV2 domain to have any safety risks.  <br /><br />
+It has not been attached to any functional domain and the LOV domain itself has no pathogenic properties.  We therefore do not expect the LOV2 domain to have any safety risks.  <br /><br/>
-It has previously been expressed in E.coli by John Christie.<br /><br />
+It has previously been expressed in E.coli by Dr John Christie.<br /><br />
-<b>References & Further Reading:</b><br />
+<b>References & Further Reading:</b><br /> <br/>
-"LOV (light, oxygen, or voltage) domains of the blue-light photoreceptor phototropin (nph1): Binding sites for the chromophore flavin mononucleotide" which can be accessed <a href=http://www.pnas.org/content/96/15/8779.short>here</a><br /><br /><br />
+"LOV (light, oxygen, or voltage) domains of the blue-light photoreceptor phototropin (nph1): Binding sites for the chromophore flavin mononucleotide" which can be accessed <a href=http://www.pnas.org/content/96/15/8779.short>here.</a><br /><br />
-"The LOV Domain Family:  Photoresponsive Signaling Modules Coupled to Diverse Output Domains" which can be accessed <a href=http://pubs.acs.org/doi/abs/10.1021/bi026978l>here</a><br /><br />
+"The LOV Domain Family:  Photoresponsive Signaling Modules Coupled to Diverse Output Domains" which can be accessed <a href=http://pubs.acs.org/doi/abs/10.1021/bi026978l>here.</a><br /><br />
-"Information on the LOV Domain by Dr John Christie" which can be accessed <a href=http://www.photobiology.info/Christie.html>here</a><Br />
+"Information on the LOV Domain" by Dr John Christie which can be accessed <a href=http://www.photobiology.info/Christie.html>here.</a><Br />
-<h1>Biobrick: LOV2 domain with 6xHIS Tag</h1>
-We have also created a version of the LOV2 biobrick with a 6xHIS tag for affinity purification.<br /><br />
+<h6><a href="#top">Return to top</a></h6>
-The use of 6xHIS tags is well established and would not give the microorganism any enhanced ability to survive outside the lab, or pose a safety risk to the public or environment.<br /><br />
+<a name="ilov"></a><h2>Biobrick: iLOV </h2>
-<b>References & Further Reading</b><br />
-"Making the most of affinity tags" which can be accessed <a href=http://www.sciencedirect.com/science/article/pii/S0167779905000843>here</a><br/>
-<h1>Biobrick: iLOV </h1>
 <b>Host:</b><br />
@@ Line 79: / Line 76: @@
 <b>Origin:</b><br />
-iLOV is a version of LOV2 that has been altered through site directed mutagenesis and DNA shuffling.
+iLOV is a version of LOV2 that has been altered through site directed mutagenesis and DNA shuffling.  <br /><br />
 <b>Function & Use</b><br />
-iLOV has the same function and uses as LOV2. As a reporter, it is advantageous over GFP derived fluorescent proteins due to its small size, ability to recover quickly from photobleaching and use in anoxic conditions. <br /><br />
+iLOV has the same function and uses as LOV2. As a reporter, it is advantageous over GFP derived fluorescent proteins due to its small size (useful if you are tagging proteins), ability to recover quickly from photobleaching and use in anoxic conditions. <br /><br />
 <b>Safety:</b><br />
@@ Line 93: / Line 90: @@
 <b>References and Further Reading:</b><br/><br />
-"The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection" which can be accessed <a href=http://www.pnas.org/content/105/50/20038.short>here</a>
+"The photoreversible fluorescent protein iLOV outperforms GFP as a reporter of plant virus infection" which can be accessed <a href=http://www.pnas.org/content/105/50/20038.short>here.</a>
+<h6><a href="#top">Return to top</a></h6>
-<h1>Biobrick: Ranaspumin (RSN2)</h1>
+<a name="rana"></a><h2>Biobrick: Ranaspumin (RSN2)</h2>
 <b>Host:</b><br />
@@ Line 115: / Line 114: @@
 <b>References & Further Reading</b><br />
-"Ranaspumin-2: Structure and Function of a Surfactant Protein from the Foam Nests of a Tropical Frog" which can be accessed <a href=http://www.sciencedirect.com/science/article/pii/S0006349509007929>here</a><br /><br />
+"Ranaspumin-2: Structure and Function of a Surfactant Protein from the Foam Nests of a Tropical Frog" which can be accessed <a href=http://www.sciencedirect.com/science/article/pii/S0006349509007929>here.</a><br /><br />
-"Biofoams and natural protein surfactants" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2954283/>here</a>
+"Biofoams and natural protein surfactants" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2954283/>here.</a>
+<h6><a href="#top">Return to top</a></h6>
-<h1>Biobrick: Ranaspumin with 6xHIS tag</h1>
+<a name="rana6"></a><h2>Biobrick: Ranaspumin with 6xHIS tag</h2>
 We have also created a version of the Ranaspumin biobrick with a 6xHIS tag. <br /><br />
 The HIS tag would not give the microorganism any enhanced ability to survive outside the lab, or pose a safety risk to the public or environment.<br /><br />
-<b>References & Further Reading</b><br/>
+<b>References & Further Reading</b><br /><br />
-"Making the most of affinity tags" which can be accessed <a href=http://www.sciencedirect.com/science/article/pii/S0167779905000843>here</a><br/>
-<h1>Biobrick: Latherin</h1>
+Information on "His-tagged proteins", which can be accessed <a href=http://www.piercenet.com/browse.cfm?fldID=1470D72F-469A-424B-90F7-2EDBCFBD33FC>here.</a><br/><br/>
+Information on "Affinity Purification", which can be accessed <a href=http://www.brunschwig-ch.com/pdf/downloads/ABT_RapidRun_Procedure.pdf?PHPSESSID=cb79f5862da62a654bb1a6aefe87c62b>here.</a>
+<h6><a href="#top">Return to top</a></h6>
+<a name="lath"></a><h2>Biobrick: Latherin</h2>
 <b>Host: </b>
@@ Line 147: / Line 152: @@
 Latherin is suspected to be involved in the allergen response to horses.  We propose that anything containing the protein should be clearly labelled as such, so that those with a horse allergy can avoid contact with it.<br /><br />
-<b>References & Further Reading:</b><br />
+<b>References & Further Reading:</b><br /><br />
 "Latherin: A Surfactant Protein of Horse Sweat and Saliva" which can be accessed <a href=http://www.plosone.org/article/info:doi%2F10.1371%2Fjournal.pone.0005726>here</a><br /><br />
-"Isolation and characterization of latherin, a surface-active protein from horse sweat" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/sites/entrez?db=pubmed&cmd=search&term=3753435>here</a><br /><br />
+"Isolation and characterization of latherin, a surface-active protein from horse sweat" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/sites/entrez?db=pubmed&cmd=search&term=3753435>here.</a><br /><br />
-"Biochemical characterization and surfactant properties of horse allergens" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/sites/entrez?db=pubmed&cmd=search&term=11358533>here</a><Br /><br />
+"Biochemical characterization and surfactant properties of horse allergens" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/sites/entrez?db=pubmed&cmd=search&term=11358533>here.</a><Br /><br />
 "Latherin and other biocompatible surfactant proteins" which can be accessed <a href=http://www.biochemsoctrans.org/bst/039/bst0391017.htm>here</a><br />
-<h1>Biobrick: Latherin with 6xHIS tag</h1>
+<h6><a href="#top">Return to top</a></h6>
+<a name="lath6"></a><h2>Biobrick: Latherin with 6xHIS tag</h2>
 We have also created a version of the Latherin biobrick with 6xHIS tag for affinity purification.<br /><br />
 The use of 6xHIS tags is well established and would not give the microorganism any enhanced ability to survive outside the lab, or pose a safety risk to the public or environment.<br /><br />
-<b>References & Further Reading</b>
+<b>References & Further Reading</b><br /><br />
-<br /><br />
-"Making the most of affinity tags" which can be accessed <a href=http://www.sciencedirect.com/science/article/pii/S0167779905000843>here</a><br/>
-<h1>Nissle 1917 Strain of <i>E.coli</i></h1>
+Information on "His-tagged proteins", which can be accessed <a href=http://www.piercenet.com/browse.cfm?fldID=1470D72F-469A-424B-90F7-2EDBCFBD33FC>here.</a><br/><br/>
+Information on "Affinity Purification", which can be accessed <a href=http://www.brunschwig-ch.com/pdf/downloads/ABT_RapidRun_Procedure.pdf?PHPSESSID=cb79f5862da62a654bb1a6aefe87c62b>here.</a>
+<h6><a href="#top">Return to top</a></h6>
+<a name="nis"></a><h2>Nissle 1917 Strain of <i>E.coli</i></h2>
 Nissle 1917 is a strain of <i>E.coli</i> that we are working with due to its ability to form biofilms.  This is a capacity that laboratory strains of <i>E.coli</i> have lost.<br /><br />
@@ Line 169: / Line 180: @@
 <b>References & Further Reading:</b><br />
-"Intestinal immunity of Escherichia coli NISSLE 1917: a safe carrier for therapeutic molecules" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/pubmed/15708311>here</a><br /><br />
+"Intestinal immunity of Escherichia coli NISSLE 1917: a safe carrier for therapeutic molecules" which can be accessed <a href=http://www.ncbi.nlm.nih.gov/pubmed/15708311>here.</a><br /><br />
+"Maintaining remission of ulcerative colitis with the probiotic Escherichia coli Nissle 1917 is as effective as with standard mesalazine" which can be accessed <a href=http://gut.bmj.com/content/53/11/1617.abstract>here.</a><br /><br />
-"Maintaining remission of ulcerative colitis with the probiotic Escherichia coli Nissle 1917 is as effective as with standard mesalazine" which can be accessed <a href=http://gut.bmj.com/content/53/11/1617.abstract>here</a><br /><br />
+"NF- B- and AP-1-Mediated Induction of Human Beta Defensin-2 in Intestinal Epithelial Cells by Escherichia coli Nissle 1917: a Novel Effect of a Probiotic Bacterium" which can be accessed <a href=http://iai.asm.org/cgi/content/abstract/72/10/5750>here.</a>
-"NF- B- and AP-1-Mediated Induction of Human Beta Defensin-2 in Intestinal Epithelial Cells by Escherichia coli Nissle 1917: a Novel Effect of a Probiotic Bacterium" which can be accessed <a href=http://iai.asm.org/cgi/content/abstract/72/10/5750>here</a>
+<h6><a href="#top">Return to top</a></h6>

Team:Glasgow/Safetybiobricks

From 2011.igem.org

Latest revision as of 04:42, 22 September 2011

Biobrick Safety

Biobrick: Phosphodiesterase (PDE)

Biobrick: LOV2

Biobrick: iLOV

Biobrick: Ranaspumin (RSN2)

Biobrick: Ranaspumin with 6xHIS tag

Biobrick: Latherin

Biobrick: Latherin with 6xHIS tag

Nissle 1917 Strain of E.coli