Team:USTC CHINA/Notebook/Protocols/HS media

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<div id="breadcrumb"><a href="https://2013.igem.org/Team:USTC_CHINA">Home</a> &gt; <a href="https://2013.igem.org/Team:USTC_CHINA/Notebook">notebook</a> &gt; <a href="https://2013.igem.org/Team:USTC_CHINA/Notebook/Protocols">protocols</a></div></div>
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<div id="breadcrumb"><a href="https://2013.igem.org/Team:USTC_CHINA">Home</a> &gt; <a href="https://2013.igem.org/Team:USTC_CHINA/Notebook">notebook</a> &gt; <a href="https://2013.igem.org/Team:USTC_CHINA/Notebook/Protocols">protocols</a>&gt; <a href="https://2013.igem.org/Team:USTC_CHINA/Notebook/Protocols/HS media">protocols HS media</a></div></div>
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<h1>Gel Extraction</h1>
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<h1>HS media</h1>
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<p>Performed with AxyPrep 96-well DNA Gel Extraction Kit (type: AP-96-GX)
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<p> 66.5 ml distilled water</br>
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Protocol
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  10 ml 10x S-base</br>
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1. Excise gel slice containing DNA fragment of interest.
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  2.5 ml 20% (w/v) glucose</br>
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2. Add 3×sample volume of Buffer DE-A.
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  5 ml 0.1% (w/v) L-tryptophan</br>
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Incubate at 75° C for 15-20 min or until gel melts completely.
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  1 ml 2% (w/v) casein</br>
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Add 0.5 × Buffer DE-A volume of Buffer DE-B.
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  5 ml 10% (w/v) yeast extract (Difco)</br>
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<img src="https://static.igem.org/mediawiki/2013/4/4d/2013igemustc-china_Protocol1.png" width="580" height="200" />  
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  10 ml 8% (w/v) arginine </br>
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3. Binding sample DNA(Centrifuge at 5,000 - 6,000×g for 5 min)
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  0.4% histidine</br>
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  autoclave all components separately</br>
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4. Add 800 μl of Buffer W2 (5,000 - 6,000×g for 1 min)
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  tryptophan solution: sterile filtration</br>
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Repeat wash with Buffer W2
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Centrifuge empty plate for 5 min at 5,000 - 6,000×g to remove residue W2.
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5. Elute with 50 μl of Eluent or Deionized Water.(5,000 - 6,000×g, 5min)
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Revision as of 15:52, 25 September 2013

HS media

66.5 ml distilled water
10 ml 10x S-base
2.5 ml 20% (w/v) glucose
5 ml 0.1% (w/v) L-tryptophan
1 ml 2% (w/v) casein
5 ml 10% (w/v) yeast extract (Difco)
10 ml 8% (w/v) arginine
0.4% histidine
autoclave all components separately
tryptophan solution: sterile filtration

notebook

Retrieved from "http://2013.igem.org/Team:USTC_CHINA/Notebook/Protocols/HS_media"