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<p align="center"><img src="https://static.igem.org/mediawiki/igem.org/d/d3/Transfo_pic_1.png" width="600px"/></p> | <p align="center"><img src="https://static.igem.org/mediawiki/igem.org/d/d3/Transfo_pic_1.png" width="600px"/></p> | ||
− | + | </div> | |
+ | <div> | ||
<h>Digestion</h> | <h>Digestion</h> | ||
<ol>1. Select restriction enzymes to digest your plasmid. Determine an appropriate reaction buffer by reading the instructions for your enzyme. | <ol>1. Select restriction enzymes to digest your plasmid. Determine an appropriate reaction buffer by reading the instructions for your enzyme. | ||
In a 1.5mL tube combine the following: | In a 1.5mL tube combine the following: | ||
− | <ul> DNA </ul> | + | <ul> - DNA </ul> |
− | <ul>Restriction Enzyme(s)</ul> | + | <ul>- Restriction Enzyme(s)</ul> |
− | <ul>Buffer </ul> | + | <ul>- Buffer </ul> |
− | <ul>dH2O up to total volume</ul> | + | <ul>- dH2O up to total volume</ul> |
<p>Mix gently by pipetting. </p></ol> | <p>Mix gently by pipetting. </p></ol> | ||
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<ol>4. To visualize the results of your digest, conduct gel electrophoresis</ol> | <ol>4. To visualize the results of your digest, conduct gel electrophoresis</ol> | ||
+ | </div> | ||
+ | <div> | ||
+ | <h>Vector Preparation</h> | ||
+ | <p>Combine the following in a PCR or Eppendorf tube:</p> | ||
+ | <ul>- 25ng Vector DNA</ul> | ||
+ | <ul>- 75ng Insert DNA</ul> | ||
+ | <ul>- Ligase Buffer (1μL/10μL reaction for 10X buffer, and 2μL/10μL reaction for 5X buffer)</ul> | ||
+ | <ul>- 0.5-1μL T4 DNA Ligase</ul> | ||
+ | <ul>- H20 to a total of 10μL</ul> | ||
+ | <p>Incubate at room temperature for 2hr, or at 16°C overnight (following the manufacturer’s instructions).</p> | ||
</div></a></li></ul></li> | </div></a></li></ul></li> |
Revision as of 14:33, 29 July 2016