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Team:British Columbia/Notebook
From 2010.igem.org
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<h3>Standard Operating Protocols (SOPS)</h3> | <h3>Standard Operating Protocols (SOPS)</h3> | ||
| + | |||
| + | <h4>Colony PCR</h4> | ||
| + | Supplies Needed: | ||
| + | <ol><li>PCR tubes</li><li>BioBrick PCR primers (G1004, G1005) or (VF2, VR)</li><li>Taq Polymerase</li><li>10x Reaction Buffer</li><li>10mM dNTPs</li><li>sdH2O</li><li>Colonies to be PCR’ed</li><li>Agar plate for indexing</li></ol> | ||
| + | |||
| + | Steps: | ||
| + | <ol><li>Make master mix of primers and other PCR components EXCEPT Taq polymerase. Keep on ice.</li> | ||
| + | <div><table align="center" width="65%" cellspacing="0" border="1" cellpadding="1"><caption> PCR Master mix</caption> | ||
| + | <tr><td>Reagent<td colspan="2">1x rxn volume (uL)<td>Master Mix</td></tr> | ||
| + | <tr><td>5x rxn buffer<td>5<td>xn<td></td></tr> | ||
| + | <tr><td>10mM dNTP<td>0.5<td>xn<td></td></tr> | ||
| + | <tr><td>sdH2O<td>9.15<td>xn<td></td></tr> | ||
| + | <tr><td>Phusion polymerase<td>0.1<td>xn<td></td></tr> | ||
| + | <tr><td>MgCl2<td>2<td>xn<td></td></tr> | ||
| + | <tr><td>DMSO - 5%<td>1.25<td>xn<td></td></tr> | ||
| + | <tr><td>gDNA<td>3<td>xn<td></td></tr> | ||
| + | <tr><td>Total<td>25</td></tr></table></div> | ||
| + | N = number of PCR tubes/samples | ||
| + | Make sure to add about 2 extra samples to account for pipetting error and 1 extra sample for water control. | ||
| + | Example: for 20 colonies, let N be: 20 (colonies) + 2 (extra)+ 1 (water) = 23 | ||
| + | <li> | ||
</div> <!-- end SubWrapper --> | </div> <!-- end SubWrapper --> | ||
Revision as of 21:54, 27 October 2010
Notebook: Need to Know
Welcome to our wiki notebook! We have organized our notebook according to sub-teams. Each page will provide you with a link to our actual notebook on OpenWetWare. To deliver the essentials here on the wiki (so you don't have to read through 6 months of experiments to get our message), we discuss the protocols, experimental outline, troubleshooting and optimization, and potential implications for iGEM.
Standard Operating Protocols (SOPS)
Colony PCR
Supplies Needed:- PCR tubes
- BioBrick PCR primers (G1004, G1005) or (VF2, VR)
- Taq Polymerase
- 10x Reaction Buffer
- 10mM dNTPs
- sdH2O
- Colonies to be PCR’ed
- Agar plate for indexing
- Make master mix of primers and other PCR components EXCEPT Taq polymerase. Keep on ice.
| Reagent | 1x rxn volume (uL) | Master Mix | |
| 5x rxn buffer | 5 | xn | |
| 10mM dNTP | 0.5 | xn | |
| sdH2O | 9.15 | xn | |
| Phusion polymerase | 0.1 | xn | |
| MgCl2 | 2 | xn | |
| DMSO - 5% | 1.25 | xn | |
| gDNA | 3 | xn | |
| Total | 25 | ||
OpenWetWare (OWW) is an effort to promote the sharing of information, know-how, and wisdom among researchers and groups who are working in biology & biological engineering. OWW hosts lab/research wikis, course wikis, protocol wikis and wiki blogs.
See our UBC OWW notebook.
