Team:SDU-Denmark/project-r

From 2010.igem.org

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(Effect)
(Biobrick assembly and sequence)
 
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We succesfully assembled the biobrick [http://partsregistry.org/Part:BBa_K343007 K343007], which consists of the TetR repressible promoter R0040 followed by the ribosome binding site B0034, the coding sequence K343003 and the terminators B0010 and B0012.<br>
We succesfully assembled the biobrick [http://partsregistry.org/Part:BBa_K343007 K343007], which consists of the TetR repressible promoter R0040 followed by the ribosome binding site B0034, the coding sequence K343003 and the terminators B0010 and B0012.<br>
This was achieved through "standard" silver assembly and confirmed via sequencing with biobrick verification forward 2 (VF2) and verification reverse (VR) primers.<br>
This was achieved through "standard" silver assembly and confirmed via sequencing with biobrick verification forward 2 (VF2) and verification reverse (VR) primers.<br>
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Sequencing results (as .ab1 files):<br>
 
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[http://2010.igem.org/wiki/images/a/ab/Sequencing_K343007.ab1| Sequencing results K343007]
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[http://partsregistry.org/cgi/sequencing/one_blast.cgi?id=8039| Sequencing results K343007]
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The sequence obtained from sequencing is identical with the theoretical sequence of the part, except for a silent mutation in the coding sequence, which does not change the amino acid sequence.
The sequence obtained from sequencing is identical with the theoretical sequence of the part, except for a silent mutation in the coding sequence, which does not change the amino acid sequence.
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The videomicroscopy indicates that bacteria carrying a plasmid with the biobrick travel father and in a more straight path than the wild type MG1655 when exposed to blue light with a wavelength around 500nm, Thereby suggesting a lowered tubling frequency. <br>
The videomicroscopy indicates that bacteria carrying a plasmid with the biobrick travel father and in a more straight path than the wild type MG1655 when exposed to blue light with a wavelength around 500nm, Thereby suggesting a lowered tubling frequency. <br>
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Computeranalysis of videos recorded of swimming baceria containing this part, indicate that these bacteria have a tendency to swim towards the source of light, when exposed to a light gradient. A trackdiagram was constructed from the gathered data, and swimming bacteria tended to orientate their movement in the direction of the light source. This could mean that blue light acts as an attractant stimulus on the SopII-HtrII-Tar complex. <br><br>
The stability of pSB1C3-K343007 is most likely <20 generations, however the stability of pSB3T5-K343007 was not determined. <br>
The stability of pSB1C3-K343007 is most likely <20 generations, however the stability of pSB3T5-K343007 was not determined. <br>
Plasmids expressing K343007 does not seem to hinder the bacterial growth in any way.<br>
Plasmids expressing K343007 does not seem to hinder the bacterial growth in any way.<br>
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We succesfully assembled the biobrick [http://partsregistry.org/Part:BBa_K343004 K343004], which consists of the TetR repressible promoter R0040 followed by the ribosome binding site B0034, the coding sequence K343000 and the terminators B0010 and B0012.<br>
We succesfully assembled the biobrick [http://partsregistry.org/Part:BBa_K343004 K343004], which consists of the TetR repressible promoter R0040 followed by the ribosome binding site B0034, the coding sequence K343000 and the terminators B0010 and B0012.<br>
This was achieved through "standard" silver assembly and confirmed via sequencing with biobrick verification forward 2 (VF2) and verification reverse (VR) primers.<br>
This was achieved through "standard" silver assembly and confirmed via sequencing with biobrick verification forward 2 (VF2) and verification reverse (VR) primers.<br>
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Sequencing results (as .ab1 files):<br>
 
<br>
<br>
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[http://2010.igem.org/wiki/images/a/ab/Sequencing_K343004.ab1| Sequencing results for K343004]
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[http://partsregistry.org/cgi/sequencing/one_blast.cgi?id=8031| Sequencing results for K343004]
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When comparing the theoretical sequence with the sequence done on the part K343004 the two sequences were identical.
When comparing the theoretical sequence with the sequence done on the part K343004 the two sequences were identical.
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The stability of pSB1C3-K343004 is most likely <20 generations, however the stability of pSB3T5-K343004 was not determined.<br>
The stability of pSB1C3-K343004 is most likely <20 generations, however the stability of pSB3T5-K343004 was not determined.<br>
The growth assay showed no significant difference between the wild type and the cells containing either pSB1C3-K343004 or pSB3K3-K343004.<br>
The growth assay showed no significant difference between the wild type and the cells containing either pSB1C3-K343004 or pSB3K3-K343004.<br>
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In a phase contrast microscope we observed that MG1655 containing pSB1C3-K343004 was longer that wild type. This support the theory that the cells are hyperflagellated and therefore stays longer in the G2-phase in the cell cycle. 
For further information, raw data and background of the assay see [http://2010.igem.org/Team:SDU-Denmark/K343004 characterization of K343004]. <br><br>
For further information, raw data and background of the assay see [http://2010.igem.org/Team:SDU-Denmark/K343004 characterization of K343004]. <br><br>
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Sequencing results (as .ab1 files):<br>
Sequencing results (as .ab1 files):<br>
<br>
<br>
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[http://2010.igem.org/wiki/images/a/ab/Sequencing_K343006.ab1| Sequencing results for K343006]
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[http://partsregistry.org/cgi/sequencing/one_blast.cgi?id=8025| Sequencing results for K343006]
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<br><br>
When comparing the theoretical sequence with the sequence done on the part K343006 the two sequences was identical.
When comparing the theoretical sequence with the sequence done on the part K343006 the two sequences was identical.
=== Effect ===
=== Effect ===
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UV-Vis spectroskopy and HPLC analysis of organic extractions of the bacteria containing K343006 didn't give conclusive results conserning the Retinal production.<br>
The stability of pSB1C3-K343006 is most likely <20 generations.<br>
The stability of pSB1C3-K343006 is most likely <20 generations.<br>
The growth assay showed no significant difference between the wild type and the cells containing pSB1C3-K343006.<br>
The growth assay showed no significant difference between the wild type and the cells containing pSB1C3-K343006.<br>

Latest revision as of 02:22, 28 October 2010