http://2013.igem.org/wiki/index.php?title=Team:BYU_Provo/Notebook/Phage_Purification/Springexp/Period1/Exp/6.19PEGPurification&feed=atom&action=historyTeam:BYU Provo/Notebook/Phage Purification/Springexp/Period1/Exp/6.19PEGPurification - Revision history2024-03-29T05:00:25ZRevision history for this page on the wikiMediaWiki 1.16.5http://2013.igem.org/wiki/index.php?title=Team:BYU_Provo/Notebook/Phage_Purification/Springexp/Period1/Exp/6.19PEGPurification&diff=103367&oldid=prevAmberb6 at 21:56, 4 September 20132013-09-04T21:56:19Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''I) Purpose'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''I) Purpose'''</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>: The first step in the phage purification process. Purify phage from bacterial debris.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>: The first step in the phage purification process. Purify phage from bacterial debris <ins class="diffchange diffchange-inline">so that we can determine the banding pattern of T7 in CsCl</ins>.</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''II) Expected Outcome'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''II) Expected Outcome'''</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: NaCl powder</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: NaCl powder</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: PEG 8000</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: PEG 8000</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>: <del class="diffchange diffchange-inline">T4 and </del>T7 bacterial lysate</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>: T7 bacterial lysate</div></td></tr>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''IV) Actual Procedure'''</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>'''IV) Actual Procedure'''</div></td></tr>
</table>Amberb6http://2013.igem.org/wiki/index.php?title=Team:BYU_Provo/Notebook/Phage_Purification/Springexp/Period1/Exp/6.19PEGPurification&diff=19910&oldid=prevDlasko10 at 22:11, 24 June 20132013-06-24T22:11:29Z<p></p>
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<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Dissolve 1.32 g of solid NaCl and let cool at 4<sup>◦</sup> C for 1 hour.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Dissolve 1.32 g of solid NaCl and let cool at 4<sup>◦</sup> C for 1 hour.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Centrifuge the suspensions at 6500 rpms (7000 g) for 10 minutes at 4<sup>◦</sup> C. Remove supernatant and put in clean flasks.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Centrifuge the suspensions at 6500 rpms (7000 g) for 10 minutes at 4<sup>◦</sup> C. Remove supernatant and put in clean flasks.</div></td></tr>
<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div>: Dissolve 3.96 g PEG 8000 and let sit at 4<sup>◦</sup> C <del class="diffchange diffchange-inline">for 1 hour</del>.</div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div>: Dissolve 3.96 g PEG 8000 and let sit at 4<sup>◦</sup> C <ins class="diffchange diffchange-inline">overnight</ins>.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Centrifuge the suspensions at 8000 rpms (10000 g) for 15 minutes at 4<sup>◦</sup> C. Discard the supernatant.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Centrifuge the suspensions at 8000 rpms (10000 g) for 15 minutes at 4<sup>◦</sup> C. Discard the supernatant.</div></td></tr>
<tr><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Turn the centrifuge bottles over to let sit for 5 minutes to drain off any remaining supernatant.</div></td><td class='diff-marker'> </td><td style="background: #eee; color:black; font-size: smaller;"><div>: Turn the centrifuge bottles over to let sit for 5 minutes to drain off any remaining supernatant.</div></td></tr>
</table>Dlasko10http://2013.igem.org/wiki/index.php?title=Team:BYU_Provo/Notebook/Phage_Purification/Springexp/Period1/Exp/6.19PEGPurification&diff=17021&oldid=prevArick6335 at 21:16, 19 June 20132013-06-19T21:16:05Z<p></p>
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<tr><td class='diff-marker'>-</td><td style="background: #ffa; color:black; font-size: smaller;"><div><font size="5"> '''6.<del class="diffchange diffchange-inline">12 </del>PEG Purification''' </font></div></td><td class='diff-marker'>+</td><td style="background: #cfc; color:black; font-size: smaller;"><div><font size="5"> '''6.<ins class="diffchange diffchange-inline">19 </ins>PEG Purification''' </font></div></td></tr>
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</table>Arick6335http://2013.igem.org/wiki/index.php?title=Team:BYU_Provo/Notebook/Phage_Purification/Springexp/Period1/Exp/6.19PEGPurification&diff=17020&oldid=prevArick6335: Created page with "{{TeamBYUProvo}} <br> {| width="100%" | colspan="3" | <font color="#333399" size="5" font face="Calibri"> '''Phage Purification May - June Notebook: Experiments'''</font> <br>..."2013-06-19T21:15:44Z<p>Created page with "{{TeamBYUProvo}} <br> {| width="100%" | colspan="3" | <font color="#333399" size="5" font face="Calibri"> '''Phage Purification May - June Notebook: Experiments'''</font> <br>..."</p>
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| colspan="3" | <font color="#333399" size="5" font face="Calibri"> '''Phage Purification May - June Notebook: Experiments'''</font><br />
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: [[Team:BYU_Provo/Phage_Purification|Overview]]<br />
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: [[Team:BYU Provo/Notebook/Phage_Purification/Winterexp|March-April]]<br />
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: [[Team:BYU Provo/Notebook/Phage_Purification/Springexp|May-June]]<br />
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: [[Team:BYU Provo/Notebook/Phage_Purification/Summerexp|July-August]]<br />
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: [[Team:BYU Provo/Notebook/Phage_Purification/Fallexp|September-October]]<br />
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<font size="5"> '''6.12 PEG Purification''' </font><br />
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'''I) Purpose'''<br />
: The first step in the phage purification process. Purify phage from bacterial debris.<br />
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'''II) Expected Outcome'''<br />
: Phage will be purified from the bacterial debris to be further purified in a CsCl gradient.<br />
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'''III) Reagants Used'''<br />
: <br />
: phage suspension buffer<br />
:: 10 mM Tris-HCl<br />
:: 100 mM NaCl<br />
:: 10 mM MgCl<sub>2</sub><br />
: DNase I<br />
: RNase A<br />
: chloroform<br />
: NaCl powder<br />
: PEG 8000<br />
: T4 and T7 bacterial lysate<br />
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'''IV) Actual Procedure'''<br />
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: Add 166.5 µL DNase and 22.5 µL RNase to T7 bacterial lysate.<br />
: Add .0904 mL chloroform to complete lysis and let the preparations sit for 30 minutes at room temperature.<br />
: Dissolve 1.32 g of solid NaCl and let cool at 4<sup>◦</sup> C for 1 hour.<br />
: Centrifuge the suspensions at 6500 rpms (7000 g) for 10 minutes at 4<sup>◦</sup> C. Remove supernatant and put in clean flasks.<br />
: Dissolve 3.96 g PEG 8000 and let sit at 4<sup>◦</sup> C for 1 hour.<br />
: Centrifuge the suspensions at 8000 rpms (10000 g) for 15 minutes at 4<sup>◦</sup> C. Discard the supernatant.<br />
: Turn the centrifuge bottles over to let sit for 5 minutes to drain off any remaining supernatant.<br />
: Suspend the pellets in .675 mL phage suspension buffer and let sit overnight at 4<sup>◦</sup> C.<br />
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'''V) Results'''<br />
: We were able to purify phage to a point that it can now be used in the CsCl gradient. We finished with phage dissolved in phage suspension buffer. We will be using this solution in a later class to further purify in a CsCl gradient.<br />
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