Composite
Part:BBa_K314104
Designed by: Tomas Huber, Justin Siegel Group: iGEM10_Washington (2010-10-08)
T7 expression cassette
Lactose (IPTG) inducible T7 promoter, BBa_K314112, protein expression insert includes f1 origin K314110, a Lac I generator K314111, a high expression promoter J23100, and the Elowitz standard RBS B0034. When ligated with a protein coding sequence at the SpeI site the scar site created will create proper spacing for protein expression.
Usage and Biology
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BBa_E0040 (GFP) was inserted between S and P using standard BioBrick assembly. The resulting constructs were grow in DH5a (constitutive promoters BBa_J23100 or BBa_J23114, as well as inducible promoter BBa_R0011) or BL21(DE3)* (T7 promoter, BBa_K314104) cells overnight at 298K in either the presence or absence of 0.5mM IPTG. The cells were harvested, washed in PBS, and then fluorescence for the whole cells was measured on a fluorometer (ex. 485, em. 525) and normalized to cell density (OD600). Cells that did not contain GFP were used as a blank. As observed, the addition of IPTG significantly increased the observed fluorescence of the cells. |
Sequence and Features
Assembly Compatibility:
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal NgoMIV site found at 126
- 1000COMPATIBLE WITH RFC[1000]
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Categories
Parameters
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