Difference between revisions of "Part:BBa K2116025"
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An esabox is an 18bp sequence to which the transcriptional regulator EsaR can bind. We placed one esabox right after the normally constitutive Anderson promoter [Bba_J23118] to create an EsaR repressible promoter. Transcription can be initiated by the specific AHL EsaR responds to [N-(3-oxo-hexanoyl)-L-homoserine lactone]. | An esabox is an 18bp sequence to which the transcriptional regulator EsaR can bind. We placed one esabox right after the normally constitutive Anderson promoter [Bba_J23118] to create an EsaR repressible promoter. Transcription can be initiated by the specific AHL EsaR responds to [N-(3-oxo-hexanoyl)-L-homoserine lactone]. | ||
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+ | When characterising this part, we used the D91G variant of EsaR, which has been documented to be more responsive to lower AHL concentrations compared to the wild type EsaR [1]. This EsaR variant was obtained through addgene, and can be found on the registry [BBa_K2116001]. | ||
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+ | Two copies of EsaR were placed under the control of individual constitutive promoters [Bba_J23118] and expressed on a medium-low copy plasmid (replication origin pBR322/rop). Based on the 2013 paper by Shong&Collins we decided to test AHL concentrations in the range of 0 to 10 000nM, since in all combinations of esabox placements tested in this paper showed saturation at 10 000nM as seen below: | ||
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+ | References: | ||
+ | [1] Shong, Jasmine and Cynthia H. Collins. "Engineering The Esar Promoter For Tunable Quorum Sensing-Dependent Gene Expression". ACS Synth. Biol. 2.10 (2013): 568-575. | ||
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Revision as of 14:17, 16 October 2016
EsaR repressible promoter
An esabox is an 18bp sequence to which the transcriptional regulator EsaR can bind. We placed one esabox right after the normally constitutive Anderson promoter [Bba_J23118] to create an EsaR repressible promoter. Transcription can be initiated by the specific AHL EsaR responds to [N-(3-oxo-hexanoyl)-L-homoserine lactone].
When characterising this part, we used the D91G variant of EsaR, which has been documented to be more responsive to lower AHL concentrations compared to the wild type EsaR [1]. This EsaR variant was obtained through addgene, and can be found on the registry [BBa_K2116001].
Two copies of EsaR were placed under the control of individual constitutive promoters [Bba_J23118] and expressed on a medium-low copy plasmid (replication origin pBR322/rop). Based on the 2013 paper by Shong&Collins we decided to test AHL concentrations in the range of 0 to 10 000nM, since in all combinations of esabox placements tested in this paper showed saturation at 10 000nM as seen below:
References: [1] Shong, Jasmine and Cynthia H. Collins. "Engineering The Esar Promoter For Tunable Quorum Sensing-Dependent Gene Expression". ACS Synth. Biol. 2.10 (2013): 568-575.
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]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]