Difference between revisions of "Part:BBa K1652000"

 
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__NOTOC__
 
__NOTOC__
 
<partinfo>BBa_K1652000 short</partinfo>
 
<partinfo>BBa_K1652000 short</partinfo>
  
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PGK1 promoter with two gal4 binding sites close to the transcription start site (TSS) that cause the promoter to be repressed in the presence of Gal4 binding protein.
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pPGK1Gx is a constitutive promoter that is able to be repressed by Gal4 transcription factor and its derivatives.
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With the GEV protein, pPGK1Gx exhibits a U-shaped dose response. GEV is a fusion protein consisting of Gal4 binding protein, human Estrogen receptor, and VP16 (a domain that recruits RNA polymerase). This promoter was created to re-purpose GEV, which is usually an activator, into a repressor. GEV activity is modulated by adding beta-estradiol to the culture. Beta-estradiol up-regulates GEV, so a higher drug concentration should increase repression. Indeed, at low concentrations of the drug, repression is seen. However, at very high concentrations, the VP16 domain on GEV actually recruits polymerase and ''activates'' the gene, although the expression level of the promoter does not reach basal levels even with an extremely high dose.
  
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<!-- Add more about the biology of this part here
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'''Dose response of pPGK1Gx to beta-estradiol'''
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===Usage and Biology===
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<html><img src="https://static.igem.org/mediawiki/2015/8/8a/Uo2015-results.jpg"></html>
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This figure shows the response of pPGK1Gx driving GFP. Concentration of beta-estradiol is given on the x-axis in nM. Fluorescence is given in arbitrary units on the y-axis. The two data series represent two biological replicates.
  
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
 
<partinfo>BBa_K1652000 SequenceAndFeatures</partinfo>
 
<partinfo>BBa_K1652000 SequenceAndFeatures</partinfo>

Revision as of 21:11, 18 September 2015

pPGK1Gx

pPGK1Gx is a constitutive promoter that is able to be repressed by Gal4 transcription factor and its derivatives.

With the GEV protein, pPGK1Gx exhibits a U-shaped dose response. GEV is a fusion protein consisting of Gal4 binding protein, human Estrogen receptor, and VP16 (a domain that recruits RNA polymerase). This promoter was created to re-purpose GEV, which is usually an activator, into a repressor. GEV activity is modulated by adding beta-estradiol to the culture. Beta-estradiol up-regulates GEV, so a higher drug concentration should increase repression. Indeed, at low concentrations of the drug, repression is seen. However, at very high concentrations, the VP16 domain on GEV actually recruits polymerase and activates the gene, although the expression level of the promoter does not reach basal levels even with an extremely high dose.

Dose response of pPGK1Gx to beta-estradiol This figure shows the response of pPGK1Gx driving GFP. Concentration of beta-estradiol is given on the x-axis in nM. Fluorescence is given in arbitrary units on the y-axis. The two data series represent two biological replicates.

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
    COMPATIBLE WITH RFC[1000]