Difference between revisions of "Part:BBa K1400003"

Revision as of 22:11, 17 October 2014

pTre(4) Single input tet responsive promoter

An improvement upon the promoter in BBa_K1164007. TThis 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 and the first two GAL4 sites are replaced with a random sequence. 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 tetr sites and not repressing sites, so increasing aTC increases activation. Estradiol has no effect beyond auto-fluorescence. Fluorescence values were give arbitrarily by the flow cytometer software.

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

@@ Line 2: / Line 2: @@
 <partinfo>BBa_K1400003 short</partinfo>
-An improvement upon the promoter in BBa_K1164007. This single input promoter has two upstream activating sequences (UAS). The third and fourth GAL4 binding site of the native pGAL1 promoter has been replaced with tetO binding sites in this version and the first and second GAL4 sites have been replaced with random sequences with identical C-G content. The Mig1 sequences that are native to the pGAL1 promoter are removed to allow transcriptional activation of the promoter in the presence of glucose in the cellular growth medium.
+An improvement upon the promoter in BBa_K1164007. TThis 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 and the first two GAL4 sites are replaced with a random sequence. 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). This is a weakly activating promoter.
+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).