Difference between revisions of "Part:BBa K2549031"
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<partinfo>BBa_K2549031 short</partinfo> | <partinfo>BBa_K2549031 short</partinfo> | ||
| − | This part is one of the response elements of our amplifier, also executing the combiner function. 8*ZF43.8 binding sites is assembled using two 4*ZF43.8 binding sites ([[Part:BBa_K2446006]]) with a biobrick scar between them. CMV ([[Part:BBa_K2549050]]) is a promotor which has a high-level constitutive expression. This part can switch off the expression of gene downstream after induced by our zinc finger-based transcription repressor. | + | This part is one of the response elements of our amplifier, also executing the combiner function. 8*ZF43.8 binding sites ([[Part:BBa_K2446013]]) is assembled using two 4*ZF43.8 binding sites ([[Part:BBa_K2446006]]) with a biobrick scar between them. CMV ([[Part:BBa_K2549050]]) is a promotor which has a high-level constitutive expression. This part can switch off the expression of gene downstream after induced by our zinc finger-based transcription repressor. |
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| + | <span class='h3bb'>Sequence and Features</span> | ||
| + | <partinfo>BBa_K2549031 SequenceAndFeatures</partinfo> | ||
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<!-- Add more about the biology of this part here --> | <!-- Add more about the biology of this part here --> | ||
===Biology=== | ===Biology=== | ||
| − | ==== | + | =====Our characterization===== |
| − | + | [[File:sTF-test.png|none|360px|thumb|'''Interaction between transcriptional repressors and their binding sites.''' A degradable EGFP (d2EGFP) is produced downstream the promoter to indicate the output strength. Experiments were conducted and analyzed as previous reported<ref>http://2017.igem.org/Team:Fudan/Demonstrate</ref>. DBD, DNA binding domain which is zinc finger in our assay. SD, silencing-form transcriptional domain; we used KRAB for the experiments in this figure. RE, responsive elements. MFI, median fluorescence intensity.]] | |
| − | + | Flow cytometry results suggest that the transcriptional repressors can effectively inhibit the promoters with responsive elements. Please visit http://2018.igem.org/Team:Fudan/Demonstration for a brief introduction of our project. | |
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| + | =====Synthetic promotor operators regulated by artificial zinc finger-based transcription factors===== | ||
| + | Khalil AS et al have reported several synthetic promotor operators which can interact with artificial zinc finger-based transcription factors with high specificity and high orthogonality<ref>A synthetic biology framework for programming eukaryotic transcription functions. Khalil AS, Lu TK, Bashor CJ, ..., Joung JK, Collins JJ. Cell, 2012 Aug;150(3):647-58 PMID: 22863014; DOI: 10.1016/j.cell.2012.05.045</ref>. | ||
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| + | [[File:ZF2.jpg|none|240px|thumb|Khalil AS et al stated:''sTFs constructed from OPEN-engineered ZFs are orthogonal to one another. sTF43-8 activated noncognate Promoter21-16 due to the fortuitous creation of a sequence that is significantly similar to the binding sequence of 43-8, when the downstream BamHI restriction site is considered.'']] | ||
<!-- Uncomment this to enable Functional Parameter display | <!-- Uncomment this to enable Functional Parameter display | ||
===Functional Parameters=== | ===Functional Parameters=== | ||
<partinfo>BBa_K2549031 parameters</partinfo> | <partinfo>BBa_K2549031 parameters</partinfo> | ||
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===References=== | ===References=== | ||
Latest revision as of 19:49, 17 October 2018
8*ZF43.8-CMV
This part is one of the response elements of our amplifier, also executing the combiner function. 8*ZF43.8 binding sites (Part:BBa_K2446013) is assembled using two 4*ZF43.8 binding sites (Part:BBa_K2446006) with a biobrick scar between them. CMV (Part:BBa_K2549050) is a promotor which has a high-level constitutive expression. This part can switch off the expression of gene downstream after induced by our zinc finger-based transcription repressor.
Sequence and Features
Assembly Compatibility:
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Biology
Our characterization
Interaction between transcriptional repressors and their binding sites. A degradable EGFP (d2EGFP) is produced downstream the promoter to indicate the output strength. Experiments were conducted and analyzed as previous reported[1]. DBD, DNA binding domain which is zinc finger in our assay. SD, silencing-form transcriptional domain; we used KRAB for the experiments in this figure. RE, responsive elements. MFI, median fluorescence intensity.
Flow cytometry results suggest that the transcriptional repressors can effectively inhibit the promoters with responsive elements. Please visit http://2018.igem.org/Team:Fudan/Demonstration for a brief introduction of our project.
Synthetic promotor operators regulated by artificial zinc finger-based transcription factors
Khalil AS et al have reported several synthetic promotor operators which can interact with artificial zinc finger-based transcription factors with high specificity and high orthogonality[2].
Khalil AS et al stated:sTFs constructed from OPEN-engineered ZFs are orthogonal to one another. sTF43-8 activated noncognate Promoter21-16 due to the fortuitous creation of a sequence that is significantly similar to the binding sequence of 43-8, when the downstream BamHI restriction site is considered.