Difference between revisions of "Part:BBa K1679018"
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<partinfo>BBa_K1679018 short</partinfo> | <partinfo>BBa_K1679018 short</partinfo> | ||
| − | This is a device containing a promoter ( | + | This is a device containing a promoter (BBa_J23109),a RNA thermometer (BBa_K115003) and a RFP coding sequence (BBa_E0010) on the pSB1C3, which can be easily transformed into E.coli. |
RNA Thermometers(RNAT) are temperature-sensing RNA sequences in 5’UTR of their mRNAs. At low temperature, RNAT folds into structure, blocking access of ribosome; at high temperature, RNAT switch from off to open conformation, increasing the efficiency of translation initiation. | RNA Thermometers(RNAT) are temperature-sensing RNA sequences in 5’UTR of their mRNAs. At low temperature, RNAT folds into structure, blocking access of ribosome; at high temperature, RNAT switch from off to open conformation, increasing the efficiency of translation initiation. | ||
When the temperature rises above 37°C, RFP would be expressed in theory. | When the temperature rises above 37°C, RFP would be expressed in theory. | ||
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===Usage and Biology=== | ===Usage and Biology=== | ||
[[File:RNAt.jpg|450px|thumb|centre]] | [[File:RNAt.jpg|450px|thumb|centre]] | ||
| − | [[File:3plateRNAT.png|450px|thumb|left| For one temperature, we spotted one strain on 3 different plates, each plate contains 9 different strains. After 48 hours, as we can see in the photo: FourU worked best under all these promoters and show great switch activity when be used cooperatively with J23101 and | + | [[File:3plateRNAT.png|450px|thumb|left| For one temperature, we spotted one strain on 3 different plates, each plate contains 9 different strains. After 48 hours, as we can see in the photo: FourU worked best under all these promoters and show great switch activity when be used cooperatively with J23101 and J23119. Our K1679018 didn't show significant difference between 37℃ and 42℃ conditions.]] |
[[File:119-003.jpg|px1500|thumb|centre|These are our K1679018 which were cultured at 28℃, 35℃, 37℃, 40℃, and 42℃ from right to left. It didn't show significant difference between these temperature in liquid LB medium.]] | [[File:119-003.jpg|px1500|thumb|centre|These are our K1679018 which were cultured at 28℃, 35℃, 37℃, 40℃, and 42℃ from right to left. It didn't show significant difference between these temperature in liquid LB medium.]] | ||
Revision as of 20:09, 18 September 2015
Promoter+RNA thermometer (PrfA) +mRFP
This is a device containing a promoter (BBa_J23109),a RNA thermometer (BBa_K115003) and a RFP coding sequence (BBa_E0010) on the pSB1C3, which can be easily transformed into E.coli. RNA Thermometers(RNAT) are temperature-sensing RNA sequences in 5’UTR of their mRNAs. At low temperature, RNAT folds into structure, blocking access of ribosome; at high temperature, RNAT switch from off to open conformation, increasing the efficiency of translation initiation. When the temperature rises above 37°C, RFP would be expressed in theory.
Usage and Biology
TUDelft-2008 has modified 3 kinds of RNAT: ROSE(BBa_K115001), FourU(BBa_ K115002), PrfA(BBa_K115003). We constructed each RNAT under the control of 3 different constitutive promoters:BBa_J23101, BBa_J23106, BBa_J23119 and use RFP as a reporter. After construction, the 9 circuits were transformed into DH5α.
According to the results of TUDelft-2008, the temperature threshold is 37℃ for FourU and PfrA, 42℃ for ROSE. Thus, we set the culture temperature to 28℃, 37℃ and 42℃ on solid LB medium while 28℃, 35℃, 37℃, 40℃, and 42℃ in liquid LB medium.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 7
Illegal NheI site found at 30 - 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 713
Illegal AgeI site found at 825 - 1000COMPATIBLE WITH RFC[1000]