Team:TU Darmstadt/protocols/Protein Expression

From 2013.igem.org

(Difference between revisions)
 
(5 intermediate revisions not shown)
Line 2: Line 2:
<style type="text/css">
<style type="text/css">
-
body
+
body  
{
{
margin:0;  
margin:0;  
Line 15: Line 15:
{
{
   background-color: white;
   background-color: white;
-
   background-image: url("/wiki/images/7/70/Hintergrund3.jpg");
+
   background-image: url("/wiki/images/5/5c/Hintergrund_Grün(ohne_Header%2C_Logo_und_Schlagschatten).jpg");
   background-attachment: scroll;
   background-attachment: scroll;
   background-position: 50% 0;
   background-position: 50% 0;
Line 26: Line 26:
{
{
     background: white;
     background: white;
-
     background-image: url('/wiki/images/2/2f/Header_(mit_Logo).jpg');
+
     background-image: url("/wiki/images/2/2f/Header_%28mit_Logo%29.jpg");
     margin: 0 auto;
     margin: 0 auto;
     height:250px ;
     height:250px ;
Line 35: Line 35:
     background-repeat: no-repeat;
     background-repeat: no-repeat;
     background-size: 100% 100% ;
     background-size: 100% 100% ;
-
     border-style: none;
+
     border-style: none;;}
-
}
+
#p-logo { display:none;}
#p-logo { display:none;}
Line 42: Line 41:
p {line-height:1.5em; margin:0 0 15px; text-align:justify; color:white;}
p {line-height:1.5em; margin:0 0 15px; text-align:justify; color:white;}
h1 {font-size:1.8em; font-weight:400; margin:0 0 12px;}
h1 {font-size:1.8em; font-weight:400; margin:0 0 12px;}
 +
#mm_icon1
#mm_icon1
{
{
position: absolute;
position: absolute;
-
top: 240px;  
+
top: 150px;  
-
left: 200px;
+
left: 30px;
}
}
#mm_icon2
#mm_icon2
{
{
-
position: relative;
+
position: absolute;
-
top: 170px;  
+
top: 150px;  
left: 350px;
left: 350px;
}
}
Line 67: Line 67:
{
{
position: absolute;
position: absolute;
-
top: 260px;  
+
top: 150px;  
-
left: 230px;
+
left: 30px;
background:white;
background:white;
filter:alpha(opacity=83); opacity:0.83;
filter:alpha(opacity=83); opacity:0.83;
Line 80: Line 80:
{
{
position: absolute;
position: absolute;
-
top: 670px;  
+
top: 150px;  
left: 350px;
left: 350px;
background:white;
background:white;
Line 89: Line 89:
border-radius:15px;
border-radius:15px;
}
}
 +
 +
#abstracticon3
#abstracticon3
Line 94: Line 96:
position: absolute;
position: absolute;
top: 150px;  
top: 150px;  
-
left: 700px;
+
left: 670px;
background:white;
background:white;
filter:alpha(opacity=83); opacity:0.83;
filter:alpha(opacity=83); opacity:0.83;
Line 102: Line 104:
border-radius:15px;
border-radius:15px;
}
}
-
 
#taskbar
#taskbar
Line 108: Line 109:
position:absolute;
position:absolute;
top:10px;
top:10px;
-
left:400px;
+
left:500px;
z-index: 5;
z-index: 5;
 +
}
 +
 +
 +
 +
 +
dl.igemTUD2013gelpicture
 +
{
 +
border: 1px solid #000;
 +
background-color: #109f71;
 +
width: 110px;
 +
text-align: center;
 +
padding: 5px 5px 5px 5px;
 +
float: right;
 +
margin: 0 0 0 0;
 +
margin-left:15px
 +
}
 +
 +
.igemTUD2013gelpicture dt
 +
{
 +
font-weight: bold;
 +
background-color: #131210;
 +
color: #959289;
 +
padding: 0 0;
 +
margin-bottom: 10px;
 +
}
 +
 +
.igemTUD2013gelpicture dd img
 +
{
 +
border: 1px solid #000;
 +
width: 100px;
 +
height: 200px;
 +
}
 +
 +
.igemTUD2013gelpicture dd
 +
{
 +
margin: 0;
 +
padding: 5px 5px 5px 5px;
 +
font-size: 100%;
 +
text-align: left;
 +
}
 +
 +
 +
 +
 +
dl.igemTUD2013gelpicture2
 +
{
 +
border: 1px solid #000;
 +
background-color: #109f71;
 +
width: 210px;
 +
text-align: center;
 +
padding: 5px 5px 5px 5px;
 +
float: right;
 +
margin: 0 0 0 0;
 +
margin-left:15px
 +
}
 +
 +
.igemTUD2013gelpicture2 dt
 +
{
 +
font-weight: bold;
 +
background-color: #131210;
 +
color: #959289;
 +
padding: 0 0;
 +
margin-bottom: 10px;
 +
}
 +
 +
.igemTUD2013gelpicture2 dd img
 +
{
 +
border: 1px solid #000;
 +
width: 200px;
 +
height: 200px;
 +
}
 +
 +
.igemTUD2013gelpicture2 dd
 +
{
 +
margin: 0;
 +
padding: 5px 5px 5px 5px;
 +
font-size: 100%;
}
}
LI.list1 {list-style: none; color:white;}
LI.list1 {list-style: none; color:white;}
Line 136: Line 214:
<img alt="Problem" src="/wiki/images/6/66/Darmstadt_green_Problem.jpg" width="100" height="30"></a>
<img alt="Problem" src="/wiki/images/6/66/Darmstadt_green_Problem.jpg" width="100" height="30"></a>
-
<a href="https://2013.igem.org/Team:TU_Darmstadt/solution">
 
-
<img alt="solution" src="/wiki/images/8/87/Darmstadt_green_Solution.jpg" width="100" height="30"></a>
 
<a href="https://2013.igem.org/Team:TU_Darmstadt/result">
<a href="https://2013.igem.org/Team:TU_Darmstadt/result">
Line 147: Line 223:
<a href="https://2013.igem.org/Team:TU_Darmstadt/team">
<a href="https://2013.igem.org/Team:TU_Darmstadt/team">
<img alt="team" src="/wiki/images/a/a4/Darmstadt_green_Team.jpg" width="70" height="30"></a>
<img alt="team" src="/wiki/images/a/a4/Darmstadt_green_Team.jpg" width="70" height="30"></a>
-
 
<br>
<br>
Line 160: Line 235:
<a href="https://2013.igem.org/Team:TU_Darmstadt/labbook">
<a href="https://2013.igem.org/Team:TU_Darmstadt/labbook">
-
<img alt="team" src="/wiki/images/f/f3/Darmstadt_green_Labbook.jpg" width="90" height="30"></a>
+
<img alt="team" src="/wiki/images/4/4c/10._Labbook_(angewählt).jpg" width="90" height="30"></a>
</div>
</div>
 +
 +
<br>
<br><br><br><br>
<br><br><br><br>
 +
<center>
 +
<a href="https://2013.igem.org/Team:TU_Darmstadt/labbook">
 +
<font size="8" color="#F0F8FF" face="Arial regular">Lab book |</font>
 +
</a>
 +
<a href="https://2013.igem.org/Team:TU_Darmstadt/materials">
 +
<font size="8" color="#F0F8FF" face="Arial regular">Materials |</font>
 +
</a>
 +
<a href="https://2013.igem.org/Team:TU_Darmstadt/protocols">
 +
<font size="8" color="#F0F8FF" face="Arial regular">Protocols</font>
 +
</a>
 +
</center>
 +
<br>
<h2><font size="6" color="#F0F8FF" face="Arial regular">Protein Expression</font></h2>
<h2><font size="6" color="#F0F8FF" face="Arial regular">Protein Expression</font></h2>
 +
<body>
-
<font size="3" color="#F0F8FF" face="Arial regular">
+
<p text-aligne:left style="margin-left:50px; margin-right:50px"><font size="4.5" color="#F0F8FF" face="Arial regular">
-
<p text-aligne:left style="margin-left:50px; margin-right:50px">
+
<B> Materials<br></B></font></p>
-
 
+
<p text-aligne:left style="margin-left:60px; margin-right:50px"><font size="3" color="#F0F8FF" face="Arial regular">
<p text-aligne:left style="margin-left:60px; margin-right:50px"><font size="3" color="#F0F8FF" face="Arial regular">
<br>
<br>
-
<B>Equipment & chemicals<br></B>
+
<B>Equipment<br></B>
<div align="left" style="margin-left:60px; margin-right:50px">
<div align="left" style="margin-left:60px; margin-right:50px">
<ul>
<ul>
<li class=list1>- Incubation shaker</li>
<li class=list1>- Incubation shaker</li>
-
<li class=list1>- <i>E. coli</i> BL21 DE3</li>
+
<li class=list1>- Photometer</li>
-
<li class=list1>- DYT media</li>
+
</ul>
 +
</div>
 +
</font>
 +
</p>
 +
 
 +
<p text-aligne:left style="margin-left:60px; margin-right:50px"><font size="3" color="#F0F8FF" face="Arial regular">
 +
<br>
 +
<B>Chemicals & consumables<br></B>
 +
<div align="left" style="margin-left:60px; margin-right:50px">
 +
<ul>
 +
<li class=list1>- E. coli BL21 DE3</li>
 +
<li class=list1>- <a href="https://2013.igem.org/Team:TU_Darmstadt/materials/Media"><font size="3" color="#F0F8FF" face="Arial regular"><b>DYT Medium</b></font></a></li>
<li class=list1>- IPTG</li>
<li class=list1>- IPTG</li>
<li class=list1>- 100 ml and 3 l flasks</li>
<li class=list1>- 100 ml and 3 l flasks</li>
-
<li class=list1>- Photometer</li>
 
<li class=list1>- ice</li>
<li class=list1>- ice</li>
</ul>
</ul>
Line 186: Line 285:
</font>
</font>
</p>
</p>
-
<br><br>
+
<br>
<p text-aligne:left style="margin-left:50px; margin-right:50px"><font size="4.5" color="#F0F8FF" face="Arial regular">
<p text-aligne:left style="margin-left:50px; margin-right:50px"><font size="4.5" color="#F0F8FF" face="Arial regular">
-
<B> Procedure<br></B></font></p>
+
<B> Procedure<br></B></font></p><br>
<p text-aligne:left style="margin-left:60px; margin-right:60px"><font size="3" color="#F0F8FF" face="Arial regular">
<p text-aligne:left style="margin-left:60px; margin-right:60px"><font size="3" color="#F0F8FF" face="Arial regular">
-
 
-
 
<div align="left" style="margin-left:30px; margin-right:50px">
<div align="left" style="margin-left:30px; margin-right:50px">
<ol>
<ol>
-
<br>
+
<li>Inoculation of 50 mL DYT medium in the 100 mL flask with <i>E. coli</i> BL21 DE3 containing <a href=http://lucerna-chem.ch/shop/558271><font size="3" color="#F0F8FF" face="Arial regular"><b> pPR-IBA2 </b></font></a>
-
<li>Inoculation of 50 mL DYT medium in the 100 mL flask with <i>E. coli</i> BL21 DE3 containing <a href=http://lucerna-chem.ch/shop/558271> pPR-IBA2 </a>
+
plasmid with the sequence for the respective part to be expressed.</li>
plasmid with the sequence for the respective part to be expressed.</li>
<li>Incubation at 180 repulsion per minute (rpm) at 30°C to an OD600= 4</li>
<li>Incubation at 180 repulsion per minute (rpm) at 30°C to an OD600= 4</li>
Line 205: Line 301:
<li>Incubation of the cell suspension over night at 180 rpm at 30°C.</li>
<li>Incubation of the cell suspension over night at 180 rpm at 30°C.</li>
</ol>
</ol>
 +
</div>
 +
</font></p>

Latest revision as of 01:59, 5 October 2013








Lab book | Materials | Protocols

Protein Expression

Materials


Equipment

  • - Incubation shaker
  • - Photometer


Chemicals & consumables

  • - E. coli BL21 DE3
  • - DYT Medium
  • - IPTG
  • - 100 ml and 3 l flasks
  • - ice


Procedure


  1. Inoculation of 50 mL DYT medium in the 100 mL flask with E. coli BL21 DE3 containing pPR-IBA2 plasmid with the sequence for the respective part to be expressed.
  2. Incubation at 180 repulsion per minute (rpm) at 30°C to an OD600= 4
  3. Transferation of the starter culture into 1 L DYT medium in a 3 L flask resulting in an OD600= 0.2
  4. Incubation to an OD600= 0.6 at 180 rpm and 30°C.
  5. Incubation for 15 minutes on ice.
  6. Induction of the proteinexpression with 20 mL of IPTG (stock conentration 1M).
  7. Incubation of the cell suspension over night at 180 rpm at 30°C.

Retrieved from "http://2013.igem.org/Team:TU_Darmstadt/protocols/Protein_Expression"