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Team:Calgary/Notebook/Protocols/Process8
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| - | TITLE=Nuclear DNA | + | TITLE=Nuclear DNA Extraction from Algae| |
BODY=<html> | BODY=<html> | ||
| - | <h4>Total DNA | + | <h4>Total DNA Isolation Protocol</h4> |
| - | + | ||
<p><h5>Reagents and Materials</h5></p> | <p><h5>Reagents and Materials</h5></p> | ||
<p><i>Extraction buffer</i> (for 40 ml total): | <p><i>Extraction buffer</i> (for 40 ml total): | ||
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</tr> | </tr> | ||
<tr> | <tr> | ||
| - | <td> | + | <td>ddH<sub>2</sub>O</td> |
<td>27 ml</td> | <td>27 ml</td> | ||
</tr> | </tr> | ||
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<p><i>Elution buffer</i> (provided in kit from Qiagen) | <p><i>Elution buffer</i> (provided in kit from Qiagen) | ||
</p> | </p> | ||
| - | |||
| + | <p><h5>Protocol</h5></p> | ||
<ol> | <ol> | ||
<li>Place 1.5 ml of algae culture into centrifuge tube.</li> | <li>Place 1.5 ml of algae culture into centrifuge tube.</li> | ||
Latest revision as of 04:14, 29 September 2011
Notebook
Post-Regionals
Journal
- Defining Objectives
- Techniques Workshop
- Bioinformatics & qRT-PCR
- Sensory Element Fishing
- Electrochem - CPR Oxidation
- Electrochem - Reporter Gene
- Pseudomonas Tools
- Microalgae Tools
- NA Viability Assays
- May - July Group Journal
Protocols
Safety
Nuclear DNA Extraction from Algae
Total DNA Isolation Protocol
Reagents and Materials
Extraction buffer (for 40 ml total):
| 1 M Tris HCl pH 7.5 | 8 ml |
| 5 M NaCl | 2 ml |
| 0.5 M EDTA | 2 ml |
| 20% SDS | 1 ml |
| ddH2O | 27 ml |
Elution buffer (provided in kit from Qiagen)
Protocol
- Place 1.5 ml of algae culture into centrifuge tube.
- Spin down cells at 5000 rpm for 1 minute.
- Pipette off supernatant.
- Add 400 µl extraction buffer.
- Incubate for 15 minutes at 50ºC, inverting tubes several times during incubation.
- Centrifuge at 13000 rpm for 5 minutes.
- Transfer supernatant to new tube.
- Add 400 µl isopropanol.
- Invert several times.
- Place on ice for 5 minutes.
- Centrifuge at 13000 rpm for 10 minutes.
- Pour off supernatant and wash pellet with 500 µL of 70% ethanol, ensuring to resuspend the pellet.
- Centrifuge at 13000 rpm for 1 minutes.
- Decant ethanol and place tubes upside down on paper towel to dry off excess ethanol.
- Dissolve pellet in 50 µl elution buffer, place tube on bench for 4 hours.
- Centrifuge at 13000 rpm for 2 minutes.
- Discard the supernatant, DNA is in the pellet.
