Team:StanfordBrownSpelman/Autoclave Media Stocks

From 2014.igem.org

(Difference between revisions)
(Created page with "<html class="no-js" lang="en"> <head> <meta charset="utf-8" /> <meta name="description" content="iGEM Wiki 2014"> <meta name="author" content="Stanford–Brown–Spelm...")
Line 43: Line 43:
<div class="row">
<div class="row">
-
   <div class="small-5 small-centered columns projecticon-cellulose-acetate"><img class="projecticon-cellulose-acetate" src="https://static.igem.org/mediawiki/2014/1/18/SBS_iGEM_2014_Waterproofing.png"/>
+
   <div class="small-5 small-centered columns projecticon-cellulose-acetate"><img class="projecticon-cellulose-acetate" src="https://static.igem.org/mediawiki/2014/6/63/SBSiGEM2014_In_The_Lab_Icon.png"/>
   </div>
   </div>
   <div id="header" class="small-8 small-centered columns">
   <div id="header" class="small-8 small-centered columns">
-
   <h3><center><a href="https://2014.igem.org/Team:StanfordBrownSpelman/Material_Waterproofing">Material Waterproofing</a></h3>
+
   <h3><center><a href="https://2014.igem.org/Team:StanfordBrownSpelman/Material_Waterproofing">Autoclave, Media &amp; Stocks</a></h3>
-
  <div class="boxedmenu"><h7><center><a href="#" id="intro">Introduction</a> ● <a href="#" id="methods">Methods</a> ● <a href="#" id="data">Results</a> ● <a href="#" id="links">References</a> ● <a href="https://2014.igem.org/Team:StanfordBrownSpelman/BioBricks#MW">BioBricks</a></h7></div>
+
<h6>
-
  <h5 id="int"><center>First approach: Paper wasp protein</h5>
+
<br /><br /><br /><br /><b>Getting Started</b>
-
<h6>Paper wasps of the genus Polistes are well known for their ability to construct nests out of transformed plant materials with paper-like properties. The most significant property of the wasp-produced paper is that it is hydrophobic and therefore waterproof. Research has identified that a protein found in the salivary glands of paper wasps is responsible for coating, strengthening, and thus waterproofing the cellulose found in plants. We collected <i>Polistes dominula</i>, an invasive European species of paper wasp, and sequence the proteins found in their saliva using a modified peptide mass fingerprinting approach. Our ultimate goal is to transform the gene coding for the wasp waterproofing protein into Saccharomyces cerevisiae so that we can produce an inherently biomimetic solution to shielding lightweight bacterial cellulose (BC) or BCOAc films from water in the environment. This project is particularly exciting because of its potential for discovery; never before have the proteins in wasp saliva been identified or applied as functional biomaterials.
+
<br />Using the Autoclave
 +
<br />Interns can’t. Ask someone with a hard badge. Typical runs take about an hour, but could be two hours if the boiler isn’t warmed up.
 +
 
 +
<br /><br />Media
 +
<br />Most of the time you'll autoclave the media after putting it together, although certain chemicals (vitamins, antibiotics) need to be added after to prevent degradation.
 +
<br /><br />LB: Use for E. coli
 +
<br />For 500mL, add:
 +
<br />5g Tryptone (or Peptone)
 +
<br />2.5g Yeast Extract
 +
<br />5g NaCl
 +
<br />7.5g Agar (if making plates)
 +
<br />Into 500ml of deionized water
 +
<br /><br />AM (Acetobacter Media): Use for <i>G. hansenii</i>
 +
<br />For 500mL, add:
 +
<br />10g Glucose
 +
<br />2.5g Tryptone (or Peptone)
 +
<br />2.5g Yeast Extract
 +
<br />1.35g Na<sub>2</sub>HPO<sub>4</sub>
 +
<br />0.75g Citric Acid
 +
<br />7.5g Agar (if making plates)
 +
<br />Into 500ml deionized water
 +
<br /><br />Antibiotic Stocks
 +
<br />We usually make our liquid stocks of antibiotics at 1000X, so that you add 1μl/mL to whatever media you are using. While the desired working concentration might change based on the plasmid, here are the stock concentrations we usually use for common antibiotics:
 +
<br />Chloramphenicol: 34mg/mL (100% EtOH)
 +
<br />Ampicillin: 100mg/mL (50% EtOH)
 +
<br />Kanamycin: 20mg/mL (H20 only)
 +
<br />Neomycin: 50mg/mL (H20 only)
 +
<br />Tetracycline: 15mg/mL (50% EtOH)
 +
<br />You'll want to make these by filter sterilizing; you cannot autoclave antibiotics. We typically store them in 1mL aliquots in a -20C or -30C freezer. Those stocks made with ethanol will not freeze. Those in water only will require thaw time.
</h6>
</h6>
-
<h5><center>Second approach: Wax ester biosynthesis</h5>
 
-
<h6>The biodegradable unmanned aerial vehicle (UAV) would be best improved if it had waterproofing capabilities. As such, various waterproofing mechanisms are under investigation for application [1]. One of the mechanisms includes the biological manipulation of the protein involved in the secretion of lipophilic wax esters from the avian uropygial gland of a pelican. Previous research has revealed that the chemical composition of the uropygial gland secretion is primarily composed of unique variations of methylhexanoic acid and fatty alcohols that react to produce wax esters. The enzymes responsible for catalyzing the esterification reaction are wax synthases. Various wax synthases have been identified across many eukaryotic and prokaryotic organisms including plants, mammals, protozoa, and bacteria. However, the current focus is bacterial and protozoan production of wax esters. Bacterial production of wax esters is most commonly associated with the <i>Acinetobacter calcoaceticus</i> bacterium and isoprenoid wax ester production in <i>Marinobacter hydrocarbonoclasticus</i> [2-3]. <i>M. hydrocarbonoclasticus</i> and <i>Euglena gracilis</i>, bacteria and protozoa respectively, were the primary focus for the synthesis of wax esters. There were two proteins that efficiently catalyzed the production of isoprenoid wax esters: wax synthase 1 and wax synthase 2. Both of these proteins can be found in <i>M. hydrocarbonoclasticus</i>. Therefore, we used molecular biology to investigate the biosynthetic production of wax esters in <i>E. coli</i> for waterproofing capabilities.  </h6>
 
   </div>
   </div>
</div>
</div>

Revision as of 02:19, 16 October 2014

Stanford–Brown–Spelman iGEM 2014 — Amberless Hell Cell

Built atop Foundation. Content &amp Development © Stanford–Brown–Spelman iGEM 2014.

Retrieved from "http://2014.igem.org/Team:StanfordBrownSpelman/Autoclave_Media_Stocks"