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Team:USTC CHINA/Notebook/Protocols/Single Digestion
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<h1>Single Digestion</h1> | <h1>Single Digestion</h1> | ||
| - | <p> | + | <p>1.Performed with Thermo Scientific FastDigest. There are four restriction endonucleases we used in our experiments: EcoRI, XbaI, SpeI, PstI.</br> |
| + | <span>Single Digestion</span></br> | ||
<table width="580" border="2"> | <table width="580" border="2"> | ||
<tr> | <tr> | ||
<td></td> | <td></td> | ||
| - | <td>Volume</td> | + | <td>Volume (μl)</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| - | <td> | + | <td>Plasmid DNA</td> |
| - | <td> | + | <td>Up to 1μg</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| - | <td> | + | <td>10×FastDigest Buffer</td> |
| - | <td> | + | <td>2</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| - | <td> | + | <td>Enzyme</td> |
| - | <td> | + | <td>1</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| - | <td> | + | <td>ddH<sub>2</sub>O</td> |
| - | <td> | + | <td>To 20</td> |
</tr> | </tr> | ||
<tr> | <tr> | ||
| - | <td> | + | <td>Total</td> |
| - | <td> | + | <td>20</td> |
</tr> | </tr> | ||
</table> | </table> | ||
| - | 2. Mix gently and spin down. | + | 2. Mix gently and spin down.</br> |
| - | 3. Incubate at 37°C in a heat block or water thermostat for 30 min. | + | 3. Incubate at 37°C in a heat block or water thermostat for 30 min.</br> |
| - | 4. Inactivate the enzyme by adding glycerol DNA loading buffer, or by heating for 5 min at 80°C(only for EcoRI) or 20min at 65°C(only for XbaI) | + | 4. Inactivate the enzyme by adding glycerol DNA loading buffer, or by heating for 5 min at 80°C(only for EcoRI) or 20min at 65°C(only for XbaI)</br> |
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Latest revision as of 08:55, 27 September 2013
