Difference between revisions of "Part:BBa K1061012"
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We conducted the functional experiment by co-tranfected the plasmid contains tTA and the plasmid contains P tight into Bosc, one kind of HEK-293 cell lines.Noted that we did not use tTA advance due to time limit(we construct the plasmid contains tTA advance later), and we will try that later,for improvement of expression control. | We conducted the functional experiment by co-tranfected the plasmid contains tTA and the plasmid contains P tight into Bosc, one kind of HEK-293 cell lines.Noted that we did not use tTA advance due to time limit(we construct the plasmid contains tTA advance later), and we will try that later,for improvement of expression control. | ||
[[Image:P_tight_functional_test.jpg|thumb|center|800px|functional assay of tTA]] | [[Image:P_tight_functional_test.jpg|thumb|center|800px|functional assay of tTA]] | ||
| − | + | After adding dox, the expression of dox is almost fully suppressed. | |
We also get the quantitive data by western blot, which will be submitted later. | We also get the quantitive data by western blot, which will be submitted later. | ||
====quantitive analysis==== | ====quantitive analysis==== | ||
Latest revision as of 02:43, 29 September 2013
tTA
tTA is a fusion protein of tetR protein from E.coli and the transcription activation domain VP16 from simplex herpes virus. It can bind to the Ptight element in response to doxycyline, and activate the expression of gene downstream of Ptight element.[1]
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Funtional experiment
We conducted the functional experiment by co-tranfected the plasmid contains tTA and the plasmid contains P tight into Bosc, one kind of HEK-293 cell lines.Noted that we did not use tTA advance due to time limit(we construct the plasmid contains tTA advance later), and we will try that later,for improvement of expression control.
After adding dox, the expression of dox is almost fully suppressed.
We also get the quantitive data by western blot, which will be submitted later.
quantitive analysis
References:
[1]http://www.clontech.com/CN/Products/Inducible_Systems/Tetracycline-Inducible_Expression/Gen2?sitex=10022:22372:US