Team:Groningen/DesignTest

From 2012.igem.org

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-
{{HeaderGroningen2012}}
+
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en">
 +
<script type="text/javascript" src="http://ajax.googleapis.com/ajax/libs/jquery/1.4.2/jquery.min.js"></script>
 +
<script src="http://ajax.googleapis.com/ajax/libs/jquery/1.4/jquery.min.js"></script>
 +
<script type="text/javascript" src="js/prototype.js"></script>
 +
<script type="text/javascript" src="js/scriptaculous.js?load=effects,builder"></script>
 +
<script type="text/javascript" src="js/lightbox.js"></script>
 +
    <script type="text/javascript" src="http://jqueryrotate.googlecode.com/svn/trunk/jQueryRotate.js"></script>
-
<html>
+
<head>
-
<head>
+
<meta http-equiv="X-UA-Compatible" content="IE=EDGE" charset="utf-8" />  
-
<style>
+
<style>
 +
div.cogbig{
 +
position:absolute;
 +
padding: 0px;
 +
margin: 0px 0px;
 +
opacity:0.4;
 +
filter:alpha(opacity=40);
 +
z-index:-1;
 +
}
 +
 +
/*Windmill*/
 +
#img1{
 +
width:120px;
 +
height:104px;
 +
position:absolute;
 +
z-index:10;
 +
left:-19px;
 +
top:-31px;
 +
}
 +
 +
/*Menu*/
 +
#navigation  {
 +
margin: 0;
 +
padding: 0 0em;
 +
height: 20px;
 +
list-style: none;
 +
display:block;
 +
font-size: 12px; 
 +
                                font-weight: 300;   
 +
color:#fff;
 +
z-index:2;
 +
}
 +
#navigation > li {
 +
float: left;
 +
height: 100%;
 +
margin-right: 18px;
 +
                                margin-left: 18px;
 +
left:100px;
 +
padding: 0 0em;
 +
}
 +
#navigation > li > a {
 +
float: left;
 +
height: 100%;
 +
color: #00;
 +
text-decoration: none;
 +
font-weight: 300;
 +
}
 +
#navigation > li > a:hover {
 +
color: Black;
 +
-
body {
 
-
font-family: Helvetica, Arial, sans-serif;
 
-
                }
+
}
-
stickerP {
+
#navigation > li.sub {
-
font-size:12pt;
+
position: relative;
-
                        line-height:12pt;
+
-
                        color:white;
+
-
}
+
-
                .sticker1 {
+
-
                        width: 200px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        padding: 10px ;
+
-
                        margin: 0px auto;
+
-
                        color: #FFF;
+
-
}
+
-
                .sticker2 {
+
-
left: 0px;
+
-
top:300px;
+
-
position:absolute;
+
-
                        width: 220px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
padding-right:10px;
+
-
padding-left:10px;
+
-
                        margin: 0 auto;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        color: #FFF;
+
-
}
+
}
-
</style>
+
#navigation > li.sub ul {
 +
width: 130px;
 +
margin: 0;
 +
padding: 0.0em 0;
 +
list-style: none;
 +
position: absolute;
 +
top: -800em;
 +
text-align:center;
 +
padding-left:13px;                                         
 +
background-color: rgba(0,0,0,0.5);
 +
}
 +
#navigation > li.sub ul li {
 +
width: 90%;
 +
background-color: rgba(0,0,0,0.9);
 +
}
 +
#navigation > li.sub ul li a {
 +
height: 100%;
 +
display: block;
 +
padding: 0.0em;
 +
color: #fff;
 +
text-decoration: none;
 +
}
 +
#navigation > li.sub ul li a:hover {
 +
color: rgb(255,103,0);
 +
font-weight: 700;
 +
}
 +
#navigation > li.sub:hover ul {
 +
top: 100px;
 +
left:-35px;
 +
}
-
<div class="sticker2">
+
#expand
-
<div class="sticker1">
+
.expand-down span
-
<stickerP ><A HREF="https://2012.igem.org/Team:Groningen/rotting" TARGET="_blank"><FONT COLOR=#ffffff>rotting of meat</FONT></A></stickerP>
+
{
-
</div>
+
font-family: Arial,Helvetica,sans-serif;
-
</div>
+
line-height: normal;
-
</body>
+
margin-bottom: 0px;
-
</head>
+
margin-top: 0px;
-
</html>
+
z-index:1;
 +
font-weight: 300;
 +
}
 +
.expand-down * {
 +
margin: 0px;
 +
padding: 0;
 +
z-index:1;
 +
}
 +
.expand-down ul {
 +
padding-top:100px;
 +
z-index:1;
 +
}
 +
.expand-down ul li {
 +
list-style-type:none;
 +
z-index:1;
 +
}
 +
.expand-down ul li a {
 +
text-decoration:none;
 +
z-index:1;
 +
}
 +
.expand-down ul li a img {
 +
width:80px;
 +
height:82px;
 +
border:none;
 +
z-index:1;
 +
}
 +
.expand-down ul li a span {
 +
//span bar under picture text
 +
z-index:1;
 +
text-align:center;
 +
font-size:14px;
 +
display: block;
 +
padding: 0.0em;
 +
color: #fff;
 +
}
 +
.expand-down ul li:hover a span {
 +
display:block;
 +
font-size:14px;
 +
z-index:1;
 +
color: rgb(255,103,0);
 +
font-weight: 700;
 +
background-color: rgba(0,0,0,0.5);
 +
}
 +
.expand-down ul li:hover a img {
 +
width:80px;
 +
height:82px;
 +
z-index:1;
 +
}
 +
.expand-down ul li:hover + li a img {
 +
width:80px;
 +
height:82px;
 +
z-index:1;
 +
}
 +
.expand-down ul li:hover + li + li a img {
 +
width:80px;
 +
height:82px;
 +
z-index:1;
 +
}
 +
/*Body*/
 +
body{
 +
background-attachment:fixed;
 +
background-color:rgb(118,28,17);
 +
text-align: justify;
 +
border: none;
 +
}
 +
#content{
 +
position: relative;
 +
top:-5px;
 +
width: 1024px;
 +
padding: 0px 0px 0px 0px;
 +
background-color:transparent;
 +
border: none;
 +
}
 +
#bodyContent{
 +
width: 1024px;
 +
position: relative;
 +
padding: 0px 0px 0px 0px;
 +
background-color:transparent;
 +
border: none;
 +
}
-
<html>
 
-
<head>
 
-
<style>
 
-
body {  
+
/*Suppress iGEM defaults*/
-
font-family: Helvetica, Arial, sans-serif;
+
a[title="Main Page"]{display: none;}
 +
.firstHeading{display: none;}
 +
.noprint{display: none;}
 +
#menubar{
 +
display: block;
 +
background-color:transparent;
 +
}
 +
#search-controls{display: none;}
 +
#contentSub{display: none;}
 +
#catlinks{display: none;}
 +
#footer-box{display: none;}
 +
#p-logo{display: block;}
 +
#top-section {
 +
display: block;
 +
position: relative;
 +
height: 98px;
 +
width: 975px;
 +
margin: -25px auto;
 +
border-left: 1px solid rgb(118,28,17);
 +
border-right: 1px solid rgb(118,28,17);
 +
border-bottom: 1px solid rgb(118,28,17);
 +
}
 +
</style>
 +
</head>
 +
 +
<body>
-
                }
 
-
GerminationP {
 
-
font-size:12pt;
 
-
                        line-height:12pt;
 
-
                        color:white;
 
-
}
 
-
                .Germination1 {
 
-
                        width: 200px;
 
-
text-align: center;
 
-
                        background: #000;
 
-
background-color: transparent;
 
-
background-color: rgba(0,0,0,0.3);
 
-
                        padding: 10px ;
 
-
                        margin: 0px auto;
 
-
                        color: #FFF;
 
-
}
 
-
                .Germination2 {
 
-
left: 550px;
 
-
top:300px;
 
-
position:absolute;
 
-
                        width: 220px;
 
-
text-align: center;
 
-
                        background: #000;
 
-
padding-right:10px;
 
-
padding-left:10px;
 
-
                        margin: 0 auto;
 
-
background-color: transparent;
 
-
background-color: rgba(0,0,0,0.3);
 
-
                        color: #FFF;
 
-
}
+
<div style="position:absolute; left: 197px; top: 30px;opacity:0.4;
-
</style>
+
filter:alpha(opacity=40);">
 +
<img  id="cog1" src="https://static.igem.org/mediawiki/2012/a/ad/Groningen2012_AD_20120802_SmallCog.png" />
 +
</div>
-
<div class="Germination2">
+
<div style="position:absolute; left: 313px; top: 30px;opacity:0.4;
-
<div class="Germination1">
+
filter:alpha(opacity=40);">
-
<GerminationP ><A HREF="https://2012.igem.org/Team:Groningen/germinationandsporulation" TARGET="_blank"><FONT COLOR=#ffffff>germination and sporulation</FONT></A></GerminationP>
+
<img  id="cog2" src="https://static.igem.org/mediawiki/2012/a/ad/Groningen2012_AD_20120802_SmallCog.png" />
</div>
</div>
-
</div>
 
-
</body>
 
-
</head>
 
-
</html>
 
-
 
+
<div style="position:absolute; left: 429px; top: 30px;opacity:0.4;
-
 
+
filter:alpha(opacity=40);">
-
<html>
+
<img  id="cog3" src="https://static.igem.org/mediawiki/2012/a/ad/Groningen2012_AD_20120802_SmallCog.png" />
-
<head>
+
-
<style>
+
-
 
+
-
body {
+
-
font-family: Helvetica, Arial, sans-serif;
+
-
 
+
-
                }
+
-
PBADmeat1P {
+
-
font-size:12pt;
+
-
                        line-height:12pt;
+
-
                        color:white;
+
-
}
+
-
                .PBADmeat11 {
+
-
                        width: 200px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        padding: 10px ;
+
-
                        margin: 0px auto;
+
-
                        color: #FFF;
+
-
}
+
-
                .PBADmeat12 {
+
-
left: 250px;
+
-
top:300px;
+
-
position:absolute;
+
-
                        width: 220px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
padding-right:10px;
+
-
padding-left:10px;
+
-
                        margin: 0 auto;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        color: #FFF;
+
-
 
+
-
}
+
-
</style>
+
-
 
+
-
<div class="PBADmeat12">
+
-
<div class="PBADmeat11">
+
-
<PBADmeat1P ><A HREF="https://2012.igem.org/Team:Groningen/identifyingbadvolatiles" TARGET="_blank"><FONT COLOR=#ffffff>dentifying bad volatiles</FONT></A></PBADmeat1>
+
</div>
</div>
-
</div>
 
-
</body>
 
-
</head>
 
-
</html>
 
-
 
+
<div style="position:absolute; left: 545px; top: 30px;opacity:0.4;
-
<html>
+
filter:alpha(opacity=40);">
-
<head>
+
<img  id="cog4" src="https://static.igem.org/mediawiki/2012/a/ad/Groningen2012_AD_20120802_SmallCog.png" />
-
<style>
+
-
 
+
-
body {
+
-
font-family: Helvetica, Arial, sans-serif;
+
-
 
+
-
                }
+
-
PBADmeat2P {
+
-
font-size:12pt;
+
-
                        line-height:12pt;
+
-
                        color:white;
+
-
}
+
-
                .PBADmeat21 {
+
-
                        width: 200px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        padding: 10px ;
+
-
                        margin: 0px auto;
+
-
                        color: #FFF;
+
-
}
+
-
                .PBADmeat22 {
+
-
left: 550px;
+
-
top:640px;
+
-
position:absolute;
+
-
                        width: 220px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
padding-right:10px;
+
-
padding-left:10px;
+
-
                        margin: 0 auto;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        color: #FFF;
+
-
 
+
-
}
+
-
</style>
+
-
 
+
-
<div class="PBADmeat22">
+
-
<div class="PBADmeat21">
+
-
<PBADmeat2P ><A HREF="https://2012.igem.org/Team:Groningen/identifyingPBADmeat" TARGET="_blank"><FONT COLOR=#ffffff>identifying PBADmeat</FONT></A></PBADmeat2P>
+
</div>
</div>
-
</div>
 
-
</body>
 
-
</head>
 
-
</html>
 
-
 
+
<div style="position:absolute; left: 661px; top: 30px;opacity:0.4;
-
<html>
+
filter:alpha(opacity=40);">
-
<head>
+
<img  id="cog5" src="https://static.igem.org/mediawiki/2012/a/ad/Groningen2012_AD_20120802_SmallCog.png" />
-
<style>
+
-
 
+
-
body {
+
-
font-family: Helvetica, Arial, sans-serif;
+
-
 
+
-
                }
+
-
PigmentsP {
+
-
font-size:12pt;
+
-
                        line-height:12pt;
+
-
                        color:white;
+
-
}
+
-
                .Pigments1 {
+
-
                        width: 200px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        padding: 10px ;
+
-
                        margin: 0px auto;
+
-
                        color: #FFF;
+
-
}
+
-
                .Pigments2 {
+
-
left: 250px;
+
-
top:680px;
+
-
position:absolute;
+
-
                        width: 220px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
padding-right:10px;
+
-
padding-left:10px;
+
-
                        margin: 0 auto;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        color: #FFF;
+
-
 
+
-
}
+
-
</style>
+
-
 
+
-
<div class="Pigments2">
+
-
<div class="Pigments1">
+
-
<PigmentsP ><A HREF="https://2012.igem.org/Team:Groningen/PBADmeat" TARGET="_blank"><FONT COLOR=#ffffff>PBADmeat</FONT></A></Pigments>
+
</div>
</div>
-
</div>
 
-
</body>
 
-
</head>
 
-
</html>
 
-
 
+
<div style="position:absolute; left: 777px; top: 30px;opacity:0.4;
-
<html>
+
filter:alpha(opacity=40);">
-
<head>
+
<img  id="cog6" src="https://static.igem.org/mediawiki/2012/a/ad/Groningen2012_AD_20120802_SmallCog.png" />
-
<style>
+
-
 
+
-
body {
+
-
font-family: Helvetica, Arial, sans-serif;
+
-
 
+
-
                }
+
-
DeathP {
+
-
font-size:12pt;
+
-
                        line-height:12pt;
+
-
                        color:white;
+
-
}
+
-
                .Death1 {
+
-
                        width: 200px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        padding: 10px ;
+
-
                        margin: 0px auto;
+
-
                        color: #FFF;
+
-
}
+
-
                .Death2 {
+
-
left: 550px;
+
-
top:680px;
+
-
position:absolute;
+
-
                        width: 220px;
+
-
text-align: center;
+
-
                        background: #000;
+
-
padding-right:10px;
+
-
padding-left:10px;
+
-
                        margin: 0 auto;
+
-
background-color: transparent;
+
-
background-color: rgba(0,0,0,0.3);
+
-
                        color: #FFF;
+
-
 
+
-
}
+
-
</style>
+
-
 
+
-
<div class="Death2">
+
-
<div class="Death1">
+
-
<DeathP ><A HREF="https://2012.igem.org/Team:Groningen/pigmentproduction" TARGET="_blank"><FONT COLOR=#ffffff>pigment production</FONT></A></DeathP>
+
</div>
</div>
-
</div>
 
-
</body>
 
-
</head>
 
-
</html>
 
 +
<div class="expand-down">
 +
<ul id="navigation" >
 +
<li class="sub" id="menuhome">
 +
 +
<a href="https://2012.igem.org/Team:Groningen" title="Home" rel="nofollow">
 +
<img src="https://static.igem.org/mediawiki/2012/6/61/Groningen2012_ID_20120808_Menu_Home.png" alt="Home"  title="Home" />
 +
<span>home</span>
 +
</a>
 +
<div class="cogbig">
 +
<img id="cogbighome" src="https://static.igem.org/mediawiki/2012/b/b6/Groningen2012_AD_20120802_BigCog.png" width="80" height="80">
 +
</div>
 +
</li>
 +
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-
==When is food rotten?==
 
-
[[File:RR_20120807_TAMCpic.jpg|680px|center]]
 
-
To make a rotting sensor, we have to have a defenition of rotten meat. For this, we used the guidelines of the European Union (2006, [http://www.imik.org/wettelijke_context/Europese_hygienerichtlijn_en_microbiologische_criteria.pdf source (in Dutch))], doing a simple Total Aerobic Microbial Count test. With this test, one can estimate the amount of colony forming units (CFU) per gram meat.
 
-
We used 70% pork, 30 % beef minced meat from our local supermarket. We incubated the meat closed from air, in portions of 1 gram at room temperature, and tested the TAMC at timepoints 0, 3, 5, 7 and 24 hours. The test has been done in triplo.
 
-
[[File:Groningen_RR_20120806_TAMC.png|500px|center]]
 
-
To see the working of our own inbuilt rotting sensor, Elbrich bravely tested the smell and look of meat for 5 hours. According to this we humans smell bad meat pretty well too. Side note: the meat has been exposed to air many times so it could be smelled. The color of the meat changed but little: it turned a bit greyer.
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==Identification of bad meat volalites by Gas Chromatography-Mass Spectometry (GC-MS)==
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===Introduction to GC-MS===
 
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With Gas Chromatography-Mass Spectrometry one can separate and identify the different volatiles present in the rotten meat. The University of Groningen has a large database of compounds available so we can identify what the GC-MS data mean. Drawback of the GC-MS is that the compounds that are identified from the rotten meat, will be destroyed during the measurements. If the GC-MS measurements succeed, it should give qualitative data, which is difficult due to the large diversity of volatiles present. We want to thank Monique Smith from Bio Organic Chemistry for her valuable help.
 
-
[[File:Groningen2012_EH_20120727_P7270578.JPG‎|500px|thumb|center|Arjan and Tom at work with the GC-MS]]
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[[File:Groningen2012_EH_20120727_P7270580.JPG‎|500px|thumb|center|Sample bottles with rotten minced meat]]
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[[File:Groningen2012_EH_20120727_P7270583.JPG‎|500px|thumb|center|All our samples, ready to be measured. The volatiles will be taken up by the syringe in the left corner (red, to the left)]]
 
-
The first step of separation in the GC is based on the fact that different compounds have different boiling points. Therefore in  complete evaporation of the compound should happen all at once and at high temperature.
 
-
Later, in the GC oven, the compounds will be condensated and different temperatures will make separation possible. The colums used for GC were HP-5 and HP-1.
 
-
Then the measurement continues with Mass Spectrometry. This is a technique to separate compounds based on their mass charge ratio. Our machine uses electron impact, ionization in vacuum and quadrapole separation. Reliability of this measurement depends on the range of the spectrum, reaction(s) with other molecules and how small the differences in molecule structure are.
 
-
===Organic solvents measurements===
 
-
The first measurements with only volatiles gave no reliable results, therefore we decided to dissolve the rotten meat into organic solvents to ensure more compounds are available for measurement.
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==Identification of BADmeat promoters by transcriptome analysis==
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===Introduction to the microarray technique===
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This technique encompasses a comparison between two conditions. These can be a knock-out strain and WT, but also the same strain on a different “diet”. In our case, ''Bacillus subtilis'' was subjected to volatiles of fresh vs. bad meat at a certain OD. We use OD 0.8-1.0, representing the late exponential phase,as this should yield much RNA. One isolates all mRNA (the transcriptome) and with reverse transcriptase, the RNA is copied into cDNA. The cDNA is tagged with a fluorescent dye, one condition red, the other green. Both conditions are then brought onto a microarray slide, where DNA of each gene of the organism of interest (in our case, ''Bacillus'') is attached in separate wells (these are called “probes”). When the cDNA is brought onto the slide, it will hybridize with the probes. After fluorescence measurement, one will see a black dot when the gene is not transcribed in both conditions, an orange/yellow one when the gene is transcribed in both, and a green or red one when just one condition causes the transcription of the gene. The ratio of red/green will be a measure for differential expression.
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{
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min-width: 350px;
 +
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 +
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 +
padding-left: 10px
 +
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 +
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 +
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 +
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 +
min-width: 350px;
 +
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 +
min-height: 200px;
 +
max-height: 200px;
 +
padding-left: 10px
 +
}
 +
table.projectmenu{
 +
background-color: transparent;
 +
}
 +
table.centertable{
 +
background-color: transparent;
 +
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 +
margin-top: 0;
 +
margin-bottom: 0;
 +
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 +
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 +
min-width: 150px;
 +
max-width: 150px;
 +
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 +
}
 +
div.bigcog{
 +
opacity:0.4;
 +
filter:alpha(opacity=40);
 +
z-index: -1;
 +
}
 +
#bigcogtopleft{
 +
position: relative;
 +
left: -10px;
 +
}
 +
#bigcogtopright{
 +
position: relative;
 +
left: 10px;
 +
}
 +
div.cogoverlay{
 +
z-index: 1;
 +
}
 +
#cogoverlaytopleft{
 +
position: relative;
 +
left: -350px;
 +
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 +
}
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 +
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 +
left: 355px;
 +
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 +
}
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#cogoverlaybottomleft{
 +
position: relative;
 +
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 +
left: 185px;
 +
}
 +
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 +
position: relative;
 +
top: -25px;
 +
left: -135px;
 +
}
 +
p.small {
 +
margin-left:0px;
 +
font-size:13pt;
 +
font-weight: normal;
 +
line-height:13pt;
 +
margin-right:0px;
 +
color:white;
 +
}
 +
p.marginChecklist{
 +
font-size:12pt;
 +
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 +
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 +
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 +
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 +
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 +
margin-left:150px;
 +
margin-right:115px;
 +
}
 +
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 +
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 +
list-style: none;
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margin-left: 70px;
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 +
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 +
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 +
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 +
.hoverbox a:hover .preview{
 +
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 +
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 +
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 +
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.hoverbox .preview{
 +
background-color: #52140C;
 +
border-color: #761C11;
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</style>
 +
</head>
-
How do we measure the transcriptome of ''Bacillus subtilis'' subjected to meat volatiles? ''Bacillus'' should be able to grow without many restrictions to maximize volatiles uptake. --> tried a few setups (also fermentors). In the end: a simple closed system, with a pot with rotten meat (rotten in fridge for 14 days), from which the air (filtered) goes through bacillus in LB medium, stirred with a magnetic stirrer. The air is pumped through the system with a simple peristaltic pump.
+
<body>
-
 
+
<br>
-
[[File:Groningen2012_EH_20120710_Volatiles_set_up.jpg|680px|center]]
+
<div class="cte">
-
{|align="center"
+
<div class="ctd">
-
|Fig. w2. Experimental set up for determining the volatiles produced by rotting meat.
+
<z1>Abstract</z1>
-
|}
+
</div>
-
 
+
</div>
-
'''Growth conditions ''Bacillus'': volatiles/oxygen depletion'''
+
<table class="centertable">
-
 
+
<tr>
-
Apart from the microarray, we’ll do this experiment again measuring the OD to see if there is a difference in growth rate for ''Bacillus''.
+
<td class="margincell" align="right">
-
 
+
<div class="bigcog" id="bigcogtopleft">
-
==Results==
+
<img src="https://static.igem.org/mediawiki/2012/b/b6/Groningen2012_AD_20120802_BigCog.png" width="150px" height="150px">
-
===Overview of overexpressed and repressed genes===
+
</div>
-
[[File:Genome2d.PNG|680px|thumb|center|Fig. w3. Part of the ''Bacillus subtilis sp 168'' genome. Genes depicted as arrows, upregulated genes are green, repressed genes pink. The color intensity is a measure for the fold of upregulation/repression]]
+
</td>
-
 
+
<td colspan="4">
-
This picture, generated with MolGen's software Genome2d, is giving an idea of the amount of up- and downregulated genes and operons in ''Bacillus subtilis'' when subjected to rotten meat volatiles.  
+
<p class="nomargin">
-
 
+
Every year, one third of global food production -1.3 billion tons of food- is thrown away, partially due to the “best before” dating system.
-
Two highly upregulated operons (upto 40 fold!), and thereby candidates for delivering our "Pbadmeat" are shown in the table below.
+
<z7>iGEM Groningen 2012</z7> seeks to provide an alternative method of assessing edibility: the <z7>Food Warden</z7>. It uses an <z7>engineered strain</z7>
-
 
+
of <i>Bacillus subtilis</i> to detect and report volatiles in spoiling meat. The introduced <z7>genetic construct</z7> uses a promoter to trigger
-
[[File:Groningen_RR_20120808_resultsmicarray.PNG|680px|center]]
+
a pigment coding gene. This promoter, <z7>identified by microarray analysis</z7>, is significantly upregulated in the presence of
-
 
+
<z7>volatiles from spoiling meat</z7>. The activity of the <z7>promoter</z7> regulates the expression of the <z7>pigment reporter</z7> and will
-
==Testing Constructs==
+
be visible to the naked eye. For safe usage of the system, spores of our engineered strain are placed into one half of a semi-permeable
-
We designed a construct for testing the different parts of our device. More information will follow, but until now, you can have a peak of the design on our office blackboard!
+
<z7>capsule</z7>, the second containing a calibrated amount of nutrients. Breaking the barrier between the two compartments allows
-
 
+
<z7>germination and growth</z7>, thereby activating the <z7>spoiling-meat sensor</z7>.
-
[[File:Groningen_RR20120807_blackboard.png|center]]
+
</p>
-
 
+
</td>
-
==Failed BioBricks==
+
<td class="margincell" align="left">
-
===BBa_K090403===
+
<div class="bigcog" id="bigcogtopright">
-
 
+
<img src="https://static.igem.org/mediawiki/2012/b/b6/Groningen2012_AD_20120802_BigCog.png" width="150px" height="150px">
-
'''Name'''  Gram-positive Shuttle Vector for Chromosomal Integration
+
</div>
-
 
+
</td>
-
'''Intended Purpose'''  Plasmid backbone for cloning in ''Bacillus subtilis''
+
</tr>
-
 
+
<tr>
-
'''Testing Protocol''' Restriction analysis (single cut)
+
<td class="margincell">
-
 
+
<div class="cogoverlay" id="cogoverlaybottomleft">
-
'''Results''' The restriction pattern showed that this biobrick is not as described in the parts registry. The cut plasmid is shorter than the expected size.
+
<img src="https://static.igem.org/mediawiki/2012/c/c2/Groningen2012_RR_20120909_rotting.png" width="100" height="100">
-
 
+
</div>
-
===BBa_I742123===
+
</td>
-
 
+
<td align="left" >
-
'''Name''' multi-host vector pTG262 converted to BioBrick vector
+
<div class="bigcog" id="bigcogbottomleft">
-
 
+
<img src="https://static.igem.org/mediawiki/2012/b/b6/Groningen2012_AD_20120802_BigCog.png" width="150px" height="150px">
-
'''Intended Purpose''' plasmid backbone for cloning in ''Bacilus subtilis''
+
</div>
-
 
+
</td>
-
'''Testing Protocol''' restriction analysis (single cut) and restriction analysis (dual cut)
+
<td align="left" >
-
 
+
<div class="cogoverlay" id="cogoverlaytopleft">
-
'''Results''' the restriction enzymes cut at illegal site(s)
+
<img src="https://static.igem.org/mediawiki/2012/1/15/Groningen2012_RR20120909_construct.png"  width="100px" height="100px">
 +
</div>
 +
</td>
 +
<td align="right" >
 +
<div class="cogoverlay" id="cogoverlaytopright">
 +
<img src="https://static.igem.org/mediawiki/2012/2/29/Groningen2012_RR20120909_badmeat.png" width="100px" height="100px">
 +
</div>
 +
</td>
 +
<td align="right">
 +
<div class="bigcog" id="bigcogbottomright">
 +
<img src="https://static.igem.org/mediawiki/2012/b/b6/Groningen2012_AD_20120802_BigCog.png" width="150px" height="150px">
 +
</div>
 +
</td>
 +
<td class="margincell">
 +
<div class="cogoverlay" id="cogoverlaybottomright">
 +
<img src="https://static.igem.org/mediawiki/2012/e/e7/Groningen2012_RR20120909_capsule.png" width="100px" height="100px">
 +
</div>
 +
</td>
 +
</tr>
 +
</table>
 +
<br>
 +
<br>
 +
<div class="cte3">
 +
<div class="ctd3">
 +
<z2 >Completed after European Regionals</z2><br><br>
 +
</div>
 +
</div>
 +
<p class="marginChecklist">
 +
<br>
 +
<br>
 +
<a href="https://2012.igem.org/Team:Groningen/in_development" target="_blank"><img src="https://static.igem.org/mediawiki/2012/d/df/IGEMGroningen2012_AD20120915_Tick.png"> Tested a construct with downregulated promoter <i>WapA</i></a><br>
 +
<a href="https://2012.igem.org/Team:Groningen/Construct" target="_blank"><img src="https://static.igem.org/mediawiki/2012/d/df/IGEMGroningen2012_AD20120915_Tick.png"> Performed enhanced construct characterization</a><br>
 +
<a href="https://2012.igem.org/Team:Groningen/Sticker" target="_blank"><img src="https://static.igem.org/mediawiki/2012/d/df/IGEMGroningen2012_AD20120915_Tick.png"> Characterized the influence of oxygen on germination and growth within the sticker</a><br>
 +
<a href="https://2012.igem.org/Team:Groningen/market_research" target="_blank"><img src="https://static.igem.org/mediawiki/2012/d/df/IGEMGroningen2012_AD20120915_Tick.png"> Expanded the scope of the market survey</a><br>
 +
<a href="https://2012.igem.org/Team:Groningen/international_cooperation" target="_blank"><img src="https://static.igem.org/mediawiki/2012/d/df/IGEMGroningen2012_AD20120915_Tick.png"> Collaborated with Cambridge 2012</a><br>
 +
<br>
 +
</p>
 +
<a name="MainAcc"></a>
 +
<div class="cte2">
 +
<div class="ctd2">
 +
<a name="MainAcc"></a><z2 >Our main accomplishments</z2><br><br>
 +
</div>
 +
</div>
 +
<br>
 +
<br>
 +
<p align=center style="color: white; font-size: 10pt;">
 +
<i>Hover your mouse over the pictures to see more!</i>
 +
</p>
 +
<br>
 +
<table class="accompli">
 +
<tr>
 +
<td class="accPic">
 +
<ul class="hoverbox">
 +
<li>
 +
<a href="#">
 +
<img src="https://static.igem.org/mediawiki/2012/c/cb/Groningen2012_RR1capsule_break.png" width=200 />
 +
<img src="https://static.igem.org/mediawiki/2012/a/ac/Groningen2012_ADStickerBig.png" class="preview" width=400 />
 +
</a>
 +
</li>
 +
</ul>
 +
</td>
 +
<td class="accPic">
 +
<ul class="hoverbox">
 +
<li>
 +
<a href="#"><img src="https://static.igem.org/mediawiki/2012/c/c2/Groningen2012_RR_20120909_rotting.png" width=150 /><img src="https://static.igem.org/mediawiki/2012/7/74/Groningen2012_ADVolatilesBig.png" class="preview" width=400 /></a>
 +
</li>
 +
</ul>
 +
</td>
 +
</tr>
 +
<tr>
 +
<td class="accCap">
 +
<p class="small">
 +
<font color=#FF6700><br><b>1.</b></font> We designed and tested the "<a class="inlink" href="https://2012.igem.org/Team:Groningen/Sticker">Sticker</a>": a capsule in which bacteria are kept inside and volatiles can go through. We used a model to get insight on how to tweak the growth of <i>Bacillus subtilis</i>.
 +
</p>
 +
</td>
 +
<td class="accCap">
 +
<p class="small">
 +
<font color=#FF6700><b>2.</b></font> We explored the definition of spoiled meat and ways to check meat spoilage. We identified <a class="inlink" href="https://2012.igem.org/Team:Groningen/volatiles">various compounds</a> present in spoiled meat.<br>
 +
</p>
 +
</td>
 +
</tr>
 +
<tr>
 +
<td class="accPic">
 +
<ul class="hoverbox">
 +
<li>
 +
<a href="#">
 +
<img src="https://static.igem.org/mediawiki/2012/2/29/Groningen2012_RR20120909_badmeat.png" width=200 />
 +
<img src="https://static.igem.org/mediawiki/2012/e/ec/Groningen2012_ADSensorBig.png" class="preview" width=400 />
 +
</a>
 +
</li>
 +
</ul>
 +
</td>
 +
<td class="accPic">
 +
<ul class="hoverbox">
 +
<li>
 +
<a href="#">
 +
<img src="https://static.igem.org/mediawiki/2012/2/24/Groningen2012_RRADPsac_transp.png" width=150 />
 +
<img src="https://static.igem.org/mediawiki/2012/c/c1/Groningen2012_AdBackboneBig.png" class="preview" width=400 />
 +
</a>
 +
</li>
 +
</ul>
 +
</td>
 +
</tr>
 +
<tr>
 +
<td class="accCap">
 +
<p class="small">
 +
<font color=#FF6700><b>3.</b></font> We identified spoiled meat <a class="inlink" href="https://2012.igem.org/Team:Groningen/Sensor">sensors</a> by transcriptome analysis.<br><br><br>
 +
</p>
 +
</td>
 +
<td class="accCap">
 +
<p class="small">
 +
<font color=#FF6700><b>4.</b></font> <a class="inlink" href="http://partsregistry.org/wiki/index.php?title=Part:BBa_K818000" target="_blank">Backbone pSac-Cm</a>: easy cloning in<br><i>B. subtilis</i>, the BioBrick way, for the first time! It's easy to check, BioBrick compatible, <i>E. coli</i> compatible, stabily inserted into the <i>B. subtilis</i> chromosome.
 +
</p>
 +
</td>
 +
</tr>
 +
<tr>
 +
<td class="accPic">
 +
<ul class="hoverbox">
 +
<li>
 +
<a href="#">
 +
<img src="https://static.igem.org/mediawiki/2012/f/f1/Groningen2012_RRADPigments_transp.png" width=150 />
 +
<img src="https://static.igem.org/mediawiki/2012/3/35/Groningen2012_ADPigmentsBig.png"  class="preview" width=400 />
 +
</a>
 +
</li>
 +
</ul>
 +
</td>
 +
<td class="accPic">
 +
<ul class="hoverbox" style="margin-left: 30px">
 +
<li>
 +
<a href="#">
 +
<img src="https://static.igem.org/mediawiki/2012/9/93/Groningen2012_ADConstruct_yellow.png" width=220 />
 +
<img src="https://static.igem.org/mediawiki/2012/2/21/Groningen2012_ADConstructBig.png" class="preview" width=400 />
 +
</a>
 +
</li>
 +
</ul>
 +
</td>
 +
</tr>
 +
<tr>
 +
<td class="accCap">
 +
<p class="small">
 +
<font color=#FF6700><br><br><b>5.</b></font> We made <a class="inlink" href="https://2012.igem.org/Team:Groningen/pigmentproduction">AmilCP and AmilGFP</a> suitable for expression in <i>Bacillus subtilis</i>.
 +
<br>
 +
These chromoproteins can be of significant value to the other <i>Bacillus subtilis</i> users in the BioBrick community.
 +
<br><br><br>
 +
</p>
 +
</td>
 +
<td class="accCap">
 +
<p class="small">
 +
<font color=#FF6700><b>6.</b></font> Most importantly: we developed a <a class="inlink" href="https://2012.igem.org/Team:Groningen/Construct">construct</a> which makes <i>Bacillus subtilis</i> sense spoiled meat and produce an output in the form of a yellow or purple pigment visible by naked eye.
 +
<br><br><br>
 +
</p>
 +
</td>
 +
</tr>
 +
</table>
 +
<br>
 +
<br>
 +
<br>
 +
</body>
 +
</html>
{{Template:SponsorsGroningen2012}}
{{Template:SponsorsGroningen2012}}

Latest revision as of 03:29, 27 October 2012





Abstract

Every year, one third of global food production -1.3 billion tons of food- is thrown away, partially due to the “best before” dating system. iGEM Groningen 2012 seeks to provide an alternative method of assessing edibility: the Food Warden. It uses an engineered strain of Bacillus subtilis to detect and report volatiles in spoiling meat. The introduced genetic construct uses a promoter to trigger a pigment coding gene. This promoter, identified by microarray analysis, is significantly upregulated in the presence of volatiles from spoiling meat. The activity of the promoter regulates the expression of the pigment reporter and will be visible to the naked eye. For safe usage of the system, spores of our engineered strain are placed into one half of a semi-permeable capsule, the second containing a calibrated amount of nutrients. Breaking the barrier between the two compartments allows germination and growth, thereby activating the spoiling-meat sensor.



Completed after European Regionals



Tested a construct with downregulated promoter WapA
Performed enhanced construct characterization
Characterized the influence of oxygen on germination and growth within the sticker
Expanded the scope of the market survey
Collaborated with Cambridge 2012

Our main accomplishments



Hover your mouse over the pictures to see more!



1. We designed and tested the "Sticker": a capsule in which bacteria are kept inside and volatiles can go through. We used a model to get insight on how to tweak the growth of Bacillus subtilis.

2. We explored the definition of spoiled meat and ways to check meat spoilage. We identified various compounds present in spoiled meat.

3. We identified spoiled meat sensors by transcriptome analysis.

4. Backbone pSac-Cm: easy cloning in
B. subtilis, the BioBrick way, for the first time! It's easy to check, BioBrick compatible, E. coli compatible, stabily inserted into the B. subtilis chromosome.



5. We made AmilCP and AmilGFP suitable for expression in Bacillus subtilis.
These chromoproteins can be of significant value to the other Bacillus subtilis users in the BioBrick community.


6. Most importantly: we developed a construct which makes Bacillus subtilis sense spoiled meat and produce an output in the form of a yellow or purple pigment visible by naked eye.





Our sponsors:

Retrieved from "http://2012.igem.org/Team:Groningen/DesignTest"