Team:UC Davis/protocols/digestion.html

From 2010.igem.org

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                 <li>Sterilized milliQ water</li>
                 <li>Sterilized milliQ water</li>
                 <li>BSA</li>
                 <li>BSA</li>
-
                 <li>NEB buffers 1,2,3,and/or 4 </li>
+
                 <li>NEB buffers 1,2,3,and/or 4 (see extra notes)</li>
                 <li>Miniprepped sample or plasmid you wish to cut</li>
                 <li>Miniprepped sample or plasmid you wish to cut</li>
                 <li>NEB digestion enzymes (EcoRI, SpeI, PstI, NheI, and/or XbaI)</li>
                 <li>NEB digestion enzymes (EcoRI, SpeI, PstI, NheI, and/or XbaI)</li>
                 </ul><p>
                 </ul><p>
<a name="procedure"><h1>Procedure</h1></a>
<a name="procedure"><h1>Procedure</h1></a>
 +
1. Add the following into a PCR tube <br />
<ul>
<ul>
-
<li>Add 22μL of milliQ water. </li>
+
<li>22μL of milliQ water </li>
 +
<li>1μL BSA </li>
 +
<li>5μL buffer x (see extra notes) </li>
 +
<li>20μL template to be cut </li>
 +
<li>1μL digestion enzyme 1 </li>
 +
<li>1μL digestion enzyme 2
 +
</ul><p>
 +
2. Run in a machine
 +
<ul>
 +
<li>37°C for 3 hours </li>
 +
<li>80°C for 20 minutes </li>
 +
<li>Keep at 4°C if to be stored </li>
</ul>
</ul>
<p>
<p>

Revision as of 17:40, 9 September 2010

Digestion

Materials

You will need:

  • Sterilized milliQ water
  • BSA
  • NEB buffers 1,2,3,and/or 4 (see extra notes)
  • Miniprepped sample or plasmid you wish to cut
  • NEB digestion enzymes (EcoRI, SpeI, PstI, NheI, and/or XbaI)

Procedure

1. Add the following into a PCR tube
  • 22μL of milliQ water
  • 1μL BSA
  • 5μL buffer x (see extra notes)
  • 20μL template to be cut
  • 1μL digestion enzyme 1
  • 1μL digestion enzyme 2

2. Run in a machine

  • 37°C for 3 hours
  • 80°C for 20 minutes
  • Keep at 4°C if to be stored

Purpose

To cut DNA at the designated places.

References

We would like to take a moment to thank all of our sponsors for their very generous donations, as we could not have done this without your help!

We would also like to thank and acknowledge:
Our Advisors
Marc Facciotti
Ilias Tagkopoulos
Technical Guidance
David Larsen
Andrew Yao
Visiting iGEMer
Jia Li of Zhejiang University (TEAM ZJU-China)
cI Promoter Screen
Drew Endy - Stanford
Thomas Schneider - NIH
Want to sponsor us? Send an email to mtfacciotti@ucdavis.edu to discuss various ways you can help! :)

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