Team:SDU-Denmark/Cambridge

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(Characterizing an existing biobrick)
 
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=Characterizing an existing biobrick=
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==Characterizing an existing biobrick==
== 2009 [https://2009.igem.org/Team:Cambridge Cambridge] brick K274210 ==
== 2009 [https://2009.igem.org/Team:Cambridge Cambridge] brick K274210 ==
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The K274210 brick was cultivated from the 2010 kit plate 3 well N6 in the PSB1A2 plasmid and transformed into E. coli strains TOP10 and MG1655.<br>
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The K274210 brick was cultivated from the 2010 kit plate 3 well N6 in the PSB1A2 plasmid and transformed into ‘’E. coli’’ strains TOP10 and MG1655.<br>
=== Characterization ===
=== Characterization ===
''' UV-vis determination of beta-carotene production'''<br>
''' UV-vis determination of beta-carotene production'''<br>
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In this experiment cells were prepared and harvested according to protocol [https://2010.igem.org/Team:SDU-Denmark/protocols#EX1.1]. This experiment was performed with four different strains of ''E. coli'': <br>
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In this experiment cells were prepared and harvested according to protocol [https://2010.igem.org/Team:SDU-Denmark/protocols#EX1.1 EX1.1]. This experiment was performed with four different strains of ''E. coli'': <br>
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Wild type TOP10<br>  
Wild type TOP10<br>  
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The measurements were performed on cells both in the exponential and stationary phase. The resulting graphs are presented below:<br>
The measurements were performed on cells both in the exponential and stationary phase. The resulting graphs are presented below:<br>
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[[Image:Team-SDU-denmarkBetacarotene samples with controles (exponential_phase).png |thumb|350px|left|UV-VIS measurement of beta-carotene concentration in four ''E. coli'' strains in Log phase]]
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[[Image:Team-SDU-denmarkBetacarotene samples with controles (exponential_phase).png |thumb|350px|left|'''Figure 1:'''UV-VIS measurement of beta-carotene concentration in four ''E. coli'' strains in Log phase]]
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[[Image:Team-SDU-denmarkBetacarotene samples with controles (stationary phase).png|thumb|350px|center|UV-VIS measurement of beta-carotene concentration in four ''E. coli'' strains in Log phase]]
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[[Image:Team-SDU-denmarkBetacarotene samples with controles (stationary phase).png|thumb|350px|center|'''Figure 2:''' UV-VIS measurement of beta-carotene concentration in four ''E. coli'' strains in Log phase]]
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In order to assess the results, a series of standard dilutions of beta-carotene were made. The concentrations were 1mM, 100 µM, 50 µM, 25 µM, 10 µM, 5 µM, 1 µM, 100 nM and 10 nM. The standard dilutions were also measured on the spectrophotometer. The resulting graphs are presented low: <br>
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In order to assess the results, a series of standard dilutions of beta-carotene were made. The concentrations were 1mM, 100 µM, 50 µM, 25 µM, 10 µM, 5 µM, 1 µM, 100 nM and 10 nM. The standard dilutions were also measured on the spectrophotometer. The resulting graphs are presented below: <br>
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[[Image:Team-SDU-denmarkStandart betacaroten curves.png |400px|thumb|center|UV-VIS standard curve of different concentrations of beta-carotene]]
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[[Image:Team-SDU-denmarkStandart betacaroten curves.png |400px|thumb|center|'''Figure 3:''' UV-VIS standard curve of different concentrations of beta-carotene]]
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When combining the graphs of the samples and the standard dilutions, it is clear that the spectres are uniform. This similarity of the spectres indicates that the cells actually do produce beta-carotene. The graphs also give a qualitative indication of the amount of beta-carotene produced by the MG1655 and the Top10 ''E. coli'' cells. The resulting graphs are presented below: <br><br>
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When combining the graphs of the samples and the standard dilutions, it is clear that the spectra are uniform. This similarity of the spectra indicates that the cells actually do produce beta-carotene. The graphs also give a qualitative indication of the amount of beta-carotene produced by the MG1655 and the Top10 ''E. coli'' cells. The resulting graphs are presented below: <br><br>
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[[Image:Image-Team-SDU-denmarkBetacarotene standarts and stationary samples.png |400px|thumb|center|The graph shows the absorbance of beta-carotene in ''E. coli'' strains MG1655 and TOP10 at different wavelengths ]]
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[[Image:Image-Team-SDU-denmarkBetacarotene standarts and stationary samples.png |400px|thumb|center|'''Figure 4:''' The graph shows the absorbance of beta-carotene in ''E. coli'' strains MG1655 and TOP10 at different wavelengths ]]
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The graph shows that the beta-carotene production of TOP10 containing pSB1A2 with constitutively active K274210 produces beta-carotene in a concentration just above 5 µM.<br>  
The graph shows that the beta-carotene production of TOP10 containing pSB1A2 with constitutively active K274210 produces beta-carotene in a concentration just above 5 µM.<br>  
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All data can be seen under  
All data can be seen under  
[https://static.igem.org/mediawiki/2010/7/7d/Team-SDU-Denmark-UV-Vis_determination_of_Carotenoids_and_polyene_chromophores_.zip Raw data]<br>
[https://static.igem.org/mediawiki/2010/7/7d/Team-SDU-Denmark-UV-Vis_determination_of_Carotenoids_and_polyene_chromophores_.zip Raw data]<br>
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The K274210 brick have also been characterized together with the K343005 and K343006 parts the data are represented in the characterization for the [https://2010.igem.org/Team:SDU-Denmark/project-p#K343005 K343005] and [https://2010.igem.org/Team:SDU-Denmark/project-p#K343006 K343006] parts.<br><br>
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The K274210 brick has also been characterized together with the K343005 and K343006 parts and the data are represented in the characterization for the [https://2010.igem.org/Team:SDU-Denmark/project-p#K343005 K343005] and [https://2010.igem.org/Team:SDU-Denmark/project-p#K343006 K343006] parts.<br><br>
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Latest revision as of 01:56, 28 October 2010