Part:BBa_K101016
Dually repressed promoter with sites for TetR and P22MNT binding
Dual tetR/p22mnt promoter, RBS, GFP, terminator. This biobrick was constructed by digesting dual promoter tet/p22mnt with EcoRI and SpeI restriction enzymes, and ligating it to XbaI and PstI digested reporter (E0240), and Eco RI and PstI digested pSB3K3. It was used for the measurement of promoter strenghth.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 42
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 732
The device has a dual promoter which is repressed by tetR and p22mnt. The input is aTc and output is fluorescence as gene for GFP is cloned downstream to the dually repressed promoter. Fluorescence was measured with different inducer amount (0.1, 0.5, 1.0 and 2 mM IPTG). Promoter activity was found to be maximum after 3 hours of induction by 100 ng/ml aTc.
Input: 50, 100, 150 and 200 ng/ml aTc
Output: Green Fluorescence (501-520 nm)
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Experiment |
Characteristic |
Value |
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Promoter Activity in DH5a Pro cells |
Maximum Fluorescence (with 100 ng/ml aTc) |
387098.4 (Normalized fluorescence at OD600 ~1.0) 3 hours post induction. |
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Minimum Fluorescence |
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2562747 (Normalized fluorescence at OD600 ~0.5) 5 minutes post induction
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Promoter Activity in TOP 10 cells |
Maximum Fluorescence |
177656.0 (Normalized fluorescence at OD600 ~1.0) 3 hours after the growth. |
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Growth of culture in LB media |
Doubling time of the recombinant cells |
~ 25 Minutes |
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Response time (after addition of IPTG) |
~20 Minutes |
Compatibility
Chassis: Device has been shown to work in
BBa_E0040, BBa_B0010, and BBa_B0010
Plasmids: Device has been shown to work in pSB3K3
| None |