Team:UCL/Labbook

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<div class="full_page">
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<div class="gap">
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</div>
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<div class="main_image"></div>
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<p class="major_title">Lab Weeks</p>
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<p class="major_title">June</p>
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<div class="gap">
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</div>
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<p class="minor_title">Week 1-3</p>
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<div class="full_page">
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<p class="body_text">
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<p class="body_text"><a href="https://2013.igem.org/Team:UCL/LabBook/Week1">Week 1</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week2"> Week 2</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week3"> Week 3</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week4"> Week 4</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week5"> Week 5</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week6"> Week 6</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week7"> Week 7</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week8"> Week 8</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week9"> Week 9</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week10"> Week 10</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week11"> Week 11</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week12"> Week 12</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week13"> Week 13</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week14"> Week 14</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week15"> Week 15</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week16"> Week 16</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week17"> Week 17</a> | <a href="https://2013.igem.org/Team:UCL/Labbook/Week18"> Week 18</a>
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No lab work
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</p>
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</p>
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</div>
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<p class="minor_title">Week 4</p>
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<div class="full_row">
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<div class="gap">
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</div>
<p class="body_text">
<p class="body_text">
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26th June - We are given safety training in the Advanced Centre for Biochemical Engineering in all relevant laboratories, as well as general procedures in case of emergency.
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Click the links above to view the corresponding Lab diary weeks
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</p>
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</p>  
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<p class="major_title">July</p>
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</div>
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<p class="minor_title">Week 5-6</p>
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<p class="body_text">
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No lab work
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</p>
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<p class="minor_title">Week 7</p>
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<p class="body_text">
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Bacterial Lab
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</p>
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15th July - The team is introduced to the laboratories which will be used during the summer for both bacterial and mammalian experiments.
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</p>
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16th July - 5X M9 salts [link to protocol], minimal agar [link to protocol], 1.4% molten agar solution [link to protocol] and 0.1M CaCl2/15% glycerol [link to protocol] were prepared for the generation of competent cells. Minimal agar plates were poured and streaked [link to streaking protocol] with W3110 Escherichia coli cells and left overnight to incubate at 37C.
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</p>
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17th July - Very little colony growth was observed from W3110 E.coli streaked plates. Plates were therefore left to incubate for a further 17 hours.
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</p>
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18th July - Sufficient colony growth allowed for the selection of a single colony from each plate. This was then inoculated in 5ul LB media [link to LB recipe] + 100ul 1M MgSO4 and left to incu-shake overnight at 37C.
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</p>
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19th July - Cultures re-suspended in new LB media and 100µl aliquots placed into individual eppendorf tubes for placement at -80C.
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</p>
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Mammalian Lab
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</p>
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17th July - Mammalian cell culture and maintenance [link to mammalian protocol] training by Mrs. Ludmilla Ruban. Passaged primary MEF (mouse embryonic fibroblast) cells. Passage 3.
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</p>
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18th July - MEF passage 4
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</p>
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19th July - MEF passage 5
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</p>
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</p>
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<p class="minor_title">Week 8</p>
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<p class="body_text">
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Bacterial Lab
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</p>
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22nd July -Transformation [link to transformation protocol] of our competent cells with plasmid YB3110 was carried out.
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</p>
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23rd July - No colony growth was observed on Ampicillin plates [link to ampicillin plate protocol] indicating no plasmid uptake. Transformation was repeated with YB3110.
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</p>
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<table>
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<tr>
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<th>Vial</th>
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<th>Ampicillin Plate</th>
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<th>Plasmid Insertion</th>
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<th>Colony Count</th>
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</tr>
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<tr>
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<td>1 (Main)</td>
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<td>Yes</td>
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<td>Yes</td>
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<td>0</td>
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</tr>
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<tr>
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<tr>
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<td>2 (Positive Control)</td>
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<td>No</td>
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<td>Yes</td>
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<td>100+</td>
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</tr>
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<tr>
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<td>3 (Negative Control)</td>
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<td>Yes</td>
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<td>No</td>
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<td>0</td>
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</tr>
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</table>
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</p>
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24th July - No colony growth from main experiment indicating no plasmid uptake. Transformation was repeated.
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</p>
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<table>
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<tr>
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<th>Vial</th>
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<th>Ampicillin Plate</th>
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<th>Plasmid Insertion</th>
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<th>Colony Count</th>
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</tr>
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<tr>
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<td>1 (Main)</td>
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<td>Yes</td>
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<td>Yes</td>
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<td>0</td>
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</tr>
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<tr>
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<tr>
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<td>2 (Positive Control)</td>
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<td>No</td>
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<td>Yes</td>
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<td>100+</td>
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</tr>
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<tr>
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<td>3 (Negative Control)</td>
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<td>Yes</td>
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<td>No</td>
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<td>0</td>
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</tr>
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</table>
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</p>
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25th July - No colony growth and hence no plasmid uptake. Transformation was repeated.
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</p>
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<table>
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<tr>
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<th>Vial</th>
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<th>Ampicillin Plate</th>
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<th>Plasmid Insertion</th>
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<th>Colony Count</th>
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</tr>
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<tr>
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<td>1 (Main)</td>
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<td>Yes</td>
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<td>Yes</td>
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<td>0</td>
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</tr>
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<tr>
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<tr>
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<td>2 (Positive Control)</td>
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<td>No</td>
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<td>Yes</td>
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<td>100+</td>
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</tr>
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<tr>
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<td>3 (Negative Control)</td>
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<td>Yes</td>
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<td>No</td>
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<td>0</td>
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</tr>
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</table>
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</p>
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26th July - Once again our cells were unsuccessful in taking up the YB3110 plasmid. We concluded that our batch of competent cells were not competent and so these will not be used in any further experiments. A new batch of competent cells are to be generated.
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</p>
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<table>
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<tr>
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<th>Vial</th>
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<th>Ampicillin Plate</th>
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<th>Plasmid Insertion</th>
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<th>Colony Count</th>
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</tr>
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<tr>
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<td>1 (Main)</td>
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<td>Yes</td>
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<td>Yes</td>
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<td>0</td>
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</tr>
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<tr>
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<tr>
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<td>2 (Positive Control)</td>
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<td>No</td>
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<td>Yes</td>
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<td>100+</td>
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</tr>
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<tr>
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<td>3 (Negative Control)</td>
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<td>Yes</td>
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<td>No</td>
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<td>0</td>
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</tr>
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</table>
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</p>
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Mammalian Lab
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</p>
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22nd July - MEF passage 6. Training ends.
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</p>
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<p class="minor_title">Week 9</p>
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<p class="body_text">
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Bacterial Lab
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</p>
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29th July - A glycerol stock of pSecTag2A from 2009 was restored. This was streaked onto 3 plates (2x LB Amp and 1x No drug), additionally four Falcons with 2ul LB were inoculated with the glycerol stock (2x LB No drug, 2X LB Amp). These were left to incubate overnight.
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</p>
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30th July - pSecTag2A Amp and No drug cultures were centrifuged (5 minutes at 4000rpm) and the pellet frozen to be used later in a miniprep. pSecTag2A plates displayed good colony growth. Colonies were picked from non-competent W3110 streaked plates and inoculated in LB ->incu-shake 37C o/n.
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</p>
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<table>
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<tr>
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<th>Vial</th>
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<th>Ampicillin Plate</th>
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<th>Plasmid Insertion</th>
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<th>Colony Count</th>
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</tr>
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<tr>
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<td>1 (Main)</td>
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<td>Yes</td>
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<td>Yes</td>
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<td>100+</td>
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</tr>
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<tr>
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<tr>
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<td>2 (Positive Control)</td>
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<td>No</td>
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<td>Yes</td>
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<td>100+</td>
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</tr>
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<tr>
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<td>3 (Negative Control)</td>
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<td>Yes</td>
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<td>No</td>
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<td>0</td>
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</tr>
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</table>
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</p>
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31st July - A new stock of competent cells was generated and stored, these were tested for competence via transformation using pSecTag2A and streaking onto amp plates -> incubate 37C o/n.
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</p>
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Ampicillin [link to Ampicillin protocol] was produced and stored. Following miniprep of pSecTag2A [link to miniprep anachem protocol] a gel [link to gel protocol] was prepared for analytical digest with HindIII [link to HindIII conditions] [insert gel image HindIII analytical digest of pSecTag2A]. 50X TAE diluted to 1X [link to dilution protocol].
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</p>
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<table>
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<tr>
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<th>Item</th>
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<th>Volume (ul)</th>
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</tr>
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<tr>
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<td>DNA pSecTag2A</td>
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<td>5</td>
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</tr>
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<tr>
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<tr>
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<td>HindIII</td>
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<td>1</td>
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</tr>
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<tr>
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<td>Buffer</td>
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<td>1</td>
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</tr>
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<tr>
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<td>BSA</td>
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<td>0.5</td>
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</tr>
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<tr>
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<td>dH20</td>
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<td>2.5</td>
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</tr>
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<tr>
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<th>Total</th>
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<td>10</td>
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</tr>
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</table>
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</p>
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<p class="major_title">August</p>
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Latest revision as of 00:25, 5 October 2013

Lab Weeks

Week 1 | Week 2 | Week 3 | Week 4 | Week 5 | Week 6 | Week 7 | Week 8 | Week 9 | Week 10 | Week 11 | Week 12 | Week 13 | Week 14 | Week 15 | Week 16 | Week 17 | Week 18

Click the links above to view the corresponding Lab diary weeks

Retrieved from "http://2013.igem.org/Team:UCL/Labbook"