Team:USTC CHINA/Notebook/Protocols/Double Digestion

From 2013.igem.org

(Difference between revisions)
(Created page with "{{USTC-China/hidden}} <html> <head> <link rel="stylesheet" type="text/css" href="https://2013.igem.org/Team:USTC_CHINA/main.css?action=raw&ctype=text/css" /> </head> <body backgro...")
Line 53: Line 53:
<div class="content" align="center">
<div class="content" align="center">
<div class="conbar1">
<div class="conbar1">
-
<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>
+
<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/Double Digestion">protocols Double Digestion</a></div></div>
<div class="conbar2">
<div class="conbar2">
<div class="leftbar" align="left">
<div class="leftbar" align="left">
<div class="bassic-bar">
<div class="bassic-bar">
-
<h1>Gel Extraction</h1>
+
<h1>Double Digestion</h1>
-
<p>Performed with AxyPrep 96-well DNA Gel Extraction Kit (type: AP-96-GX)
+
<p>1.Performed with Thermo Scientific FastDigest. There are four restriction endonucleases we used in our experiments: EcoRI, XbaI, SpeI, PstI.
-
Protocol
+
-
1. Excise gel slice containing DNA fragment of interest.
+
-
2. Add 3×sample volume of Buffer DE-A.
+
-
Incubate at 75° C for 15-20 min or until gel melts completely.
+
-
Add 0.5 × Buffer DE-A volume of Buffer DE-B.
+
-
<img src="https://static.igem.org/mediawiki/2013/4/4d/2013igemustc-china_Protocol1.png" width="580" height="200" />
+
-
3. Binding sample DNA(Centrifuge at 5,000 - 6,000×g for 5 min)
+
-
+
-
4. Add 800 μl of Buffer W2 (5,000 - 6,000×g for 1 min)
+
-
Repeat wash with Buffer W2
+
-
Centrifuge empty plate for 5 min at 5,000 - 6,000×g to remove residue W2.
+
-
+
-
5. Elute with 50 μl of Eluent or Deionized Water.(5,000 - 6,000×g, 5min)
+
-
+
-
 
+
</p></div>
</p></div>
</div>
</div>

Revision as of 16:31, 25 September 2013

Double Digestion

1.Performed with Thermo Scientific FastDigest. There are four restriction endonucleases we used in our experiments: EcoRI, XbaI, SpeI, PstI.

notebook

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