Difference between revisions of "Part:BBa K1400003"

Latest revision as of 02:58, 18 October 2014

pTre(4) Single input tet responsive promoter

An improvement upon the promoter in BBa_K1164007. This single input promoter has four upstream activating sequences (UAS). The four UAS sites are tetO binding sites that can bind to the tetracycline responsive activator protein, rtTA (reverse tetracycline-controlled transactivator), to induce transcription. The third and fourth GAL4 binding sites of the native pGAL1 promoter are replaced with tetO sites in this version. The Mig1 sequences that are native to the pGAL1 promoter are replaced with two tetO sites. In cells expressing rtTA, this promoter can be used to drive transcription of a downstream gene by the addition of aTc (anhydrotetracycline).

It was designed to have similar expression to our modified gal promoter (BBA_K1400002).

Figure 1: Characterization of pTRE via dual drug induction. pTRE has 4 activating tetO sites and no repressing sites, so increasing aTC increases activation. Estradiol has no effect beyond auto-fluorescence. Fluorescence values were given arbitrarily by the flow cytometer software. The fusion protein, rtTA (reverse tetracycline-controlled transactivator), is the activator and is expressed with the weak constitutive promoter, pMRP7.

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
INCOMPATIBLE WITH RFC[25]
Illegal AgeI site found at 35
1000
INCOMPATIBLE WITH RFC[1000]
Illegal BsaI.rc site found at 166

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 <partinfo>BBa_K1400003 short</partinfo>
-An improvement upon the promoter in BBa_K1164007. A single tet responsive promoter created by removing the 2 most distal (from TATA box) gal4 sites, and replacing the last 2 gal4 sites in the gal promoter with tet sites. THe two mig1 sites were replaced as well. It was designed to have similar expression to our modified gal promoter (BBA_K1400002).
+An improvement upon the promoter in <html><a href="https://parts.igem.org/Part:BBa_K1164007">BBa_K1164007</a></html>. This single input promoter has four upstream activating sequences (UAS). The four UAS sites are tetO binding sites that can bind to the tetracycline responsive activator protein, rtTA (reverse tetracycline-controlled transactivator), to induce transcription. The third and fourth GAL4 binding sites of the native pGAL1 promoter are replaced with tetO sites in this version. The Mig1 sequences that are native to the pGAL1 promoter are replaced with two tetO sites.
+In cells expressing rtTA, this promoter can be used to drive transcription of a downstream gene by the addition of aTc (anhydrotetracycline).
+It was designed to have similar expression to our modified gal promoter (<html><a href="https://parts.igem.org/wiki/index.php?title=Part:BBa_K1400002">BBA_K1400002</a></html>).
+<html><img style="width:65%;" src="https://static.igem.org/mediawiki/2014/d/d3/PTRE_4_sites.png" /></html>
+Figure 1: Characterization of pTRE via dual drug induction. pTRE has 4 activating tetO sites and no repressing sites, so increasing aTC increases activation. Estradiol has no effect beyond auto-fluorescence. Fluorescence values were given arbitrarily by the flow cytometer software. The fusion protein, rtTA (reverse tetracycline-controlled transactivator), is the activator and is expressed with the weak constitutive promoter, pMRP7.
 <!-- Add more about the biology of this part here