Team:NTNU-Trondheim/Notebook/October

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<p style="text-align:center; color:black; "> October</p> </div>
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[https://2013.igem.org/Team:NTNU-Trondheim/TransformedBricks Transformed BioBricks]
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<p style="text-align:center; color:black; "> Wednesday 02.10.2013</p> </div>
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<p style="text-align:center; color:black; ">SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct</p> </div>
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<p>In order to determine if Protein G is produced in the S3-2B sample and wheater a GFP-RFP dimer is produced in the ER1 sample, a SDS-PAGE was performed.
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Liquid cell culture with ER1 (tat_GFP_RFP construct), S3-2B (tat_ProteinG construct) and wildtype ER2566 ''E.coli'' cells was centrifuged and the pallet was mixed with SDS loading buffer. This mix was then heated at 95 &deg;C for 15 minutes. The SDS-PAGE results can be viewed in the figure below
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<div class="col4" style="background-color:white;><a href="https://static.igem.org/mediawiki/2013/8/8e/Bakteri_SDS_gel_2.jpg"> <img src="https://static.igem.org/mediawiki/2013/8/8e/Bakteri_SDS_gel_2.jpg" width="303">
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====<center>Wednesday 02.10.2013</center>====
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<p style="text-align:center; color:black; "> <b>Figure 1:</b> SDS-PAGE of bacterial samples. From the left: wild type ER2566, tat_GFP_RFP and tat_ProteinG</p> </div>
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=====SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct=====
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In order to determine if Protein G is produced in the S3-2B sample and wheater a GFP-RFP dimer is produced in the ER1 sample, a SDS-PAGE was performed.
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Liquid cell culture with ER1 (tat_GFP_RFP construct), S3-2B (tat_ProteinG construct) and wildtype ER2566 ''E.coli'' cells was centrifuged and the pallet was mixed with SDS loading buffer. This mix was then heated at 95 &deg;C for 15 minutes. The SDS-PAGE results can be viewed in the figure below
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There is evidently produced Protein G in the S3-2B sample as there is an additional band at the expected size of about 60 kDa. There is also an additional band at the ER1 sample of about 28-30 kDa. This half the expected size of about 60 kDa. This indicates that only RFP is produced in the bacterias.<br></p>
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<p style="text-align:center; color:black; "> Figure 1.</p> </div>
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<center>[[file:SDS-PAGE_bacterias.jpg|400px]]</center>
 
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<center>Figure: SDS-PAGE of bacterial samples. From the left: wild type ER2566, tat_GFP_RFP and tat_ProteinG</center>
 
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<p style="text-align:center; color:black; ">SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct</p> </div>
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<br><br>
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<p>A new test on the Pm/XylS promoter was performed with the same conditions as earlier.<br></p>
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<div class ="row-end"> </div>
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There is evidently produced Protein G in the S3-2B sample as there is an additional band at the expected size of about .... kDa. There is also an additional band at the ER1 sample of about ..... kDa. This half the expected size of about 60 kDa. This indicates that only RFP is produced in the bacterias.<br></p>
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<p style="text-align:center; color:black; "> Wednesday 02 - Friday 04.10.2013</p> </div>
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As the first attempt to send in the tat_GFP_RFP construct to iGEM Headquarters failed, we redid the cloning as we did 26-28.09.2013 and sent in a new construct.
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Latest revision as of 22:43, 4 October 2013

Trondheim iGEM 2013

header
Mercury
October

Wednesday 02.10.2013

SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct



In order to determine if Protein G is produced in the S3-2B sample and wheater a GFP-RFP dimer is produced in the ER1 sample, a SDS-PAGE was performed. Liquid cell culture with ER1 (tat_GFP_RFP construct), S3-2B (tat_ProteinG construct) and wildtype ER2566 ''E.coli'' cells was centrifuged and the pallet was mixed with SDS loading buffer. This mix was then heated at 95 °C for 15 minutes. The SDS-PAGE results can be viewed in the figure below

Figure 1: SDS-PAGE of bacterial samples. From the left: wild type ER2566, tat_GFP_RFP and tat_ProteinG

There is evidently produced Protein G in the S3-2B sample as there is an additional band at the expected size of about 60 kDa. There is also an additional band at the ER1 sample of about 28-30 kDa. This half the expected size of about 60 kDa. This indicates that only RFP is produced in the bacterias.

SDS-PAGE of bacterial samples with the tat_GFP_RFP (ER1) and tat_ProteinG construct



A new test on the Pm/XylS promoter was performed with the same conditions as earlier.

Wednesday 02 - Friday 04.10.2013

As the first attempt to send in the tat_GFP_RFP construct to iGEM Headquarters failed, we redid the cloning as we did 26-28.09.2013 and sent in a new construct.


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