Team:StanfordBrownSpelman/Autoclave Media Stocks
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- | <h3><center><a href="https://2014.igem.org/Team:StanfordBrownSpelman/Material_Waterproofing"> | + | <h3><center><a href="https://2014.igem.org/Team:StanfordBrownSpelman/Material_Waterproofing">Autoclave, Media & Stocks</a></h3> |
- | + | <h6> | |
- | + | <br /><br /><br /><br /><b>Getting Started</b> | |
- | + | <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. | ||
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Revision as of 02:19, 16 October 2014