Difference between revisions of "Part:BBa K584004"
 
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<partinfo>BBa_K584004 short</partinfo>
 
<partinfo>BBa_K584004 short</partinfo>
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This part was designed to test the functionality of the promotor region. We cloned the HybB promoter via PCR using the [https://parts.igem.org/Part:BBa_K410000 K410000] as a template using the primers below -
 
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<font color=#006600>hyb-FW: CCGGAATTCGCGGCCGCTTCTAGAGCGCCGCTATGGACTGGATAAAG </font>
 
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<font color=#006600>hyb-RV: AAAACTGCAGCGGCCGCTACTAGTATGCTACTTAACCCCATGGTGG  </font>   
 
  
 
<u><b>Characterization</b></u>
 
<u><b>Characterization</b></u>
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<p align="justify">To test the usefulness of the cold shock-inducible promoter J45503 in our 2011 iGEM project, we fused the promoter to a GFP reporter, and assayed the promoter’s activity after a temperature shift from 37°C to 25°C or 4°C. We tested this activity both in a TOP10F’ (figure 1) as well as a MG1655 (figure 2) E.coli strain background. For more information on E.coli strain descriptions, we recommend the following web site: [http://openwetware.org/wiki/E._coli_genotypes E.Coli_Genotypes].</p>
 
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[[Image:HybB-GFP_TOP10.jpg|thumb|center|Figure1]]
 
  
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[[Image:HybB-GFP_MG1655.jpg|thumb|center|Figure2]]
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<p align="justify">The <b>[http://2011.igem.org/Team:KULeuven K.U.Leuven 2011 iGEM Team]</b> has <b><u>extensively characterized</u></b> the <b><u>existing Biobrick part</u></b> [https://parts.igem.org/wiki/index.php?title=Part:BBa_J45503 hybB Cold Shock Promoter(BBa_J45503)].</p>
  
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<p align="justify">We can see clearly that transferring cells (both TOP10F’ and MG1655) to lower temperatures (4°C and even 25°C) results in a growth arrest between the 1 and 4 hour time points of our experiment (Figures 1A and 2A). We see that promoter activity is induced when cells are transferred to 25°C and even when they are put in and ice bath (4°C) (Figures 1B and 2B). Unfortunately, however, cells that are kept at 37°C also display an increase in promoter activity, indicating leakiness in the system.</p>
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The detailed qualitative and quantitative <u><b>[https://parts.igem.org/Part:BBa_J45503:Experience#User_Reviews review]</b></u> has been added into the experience page of the BBa_J45503 hybB Cold Shock Promoter.
  
 
<!-- Add more about the biology of this part here
 
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Latest revision as of 10:29, 21 September 2011

HybB promoter + GFP generator

Characterization

The [http://2011.igem.org/Team:KULeuven K.U.Leuven 2011 iGEM Team] has extensively characterized the existing Biobrick part hybB Cold Shock Promoter(BBa_J45503).

The detailed qualitative and quantitative review has been added into the experience page of the BBa_J45503 hybB Cold Shock Promoter.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1064