Difference between revisions of "Part:BBa K2995000"
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Constitutive promoter for expression in E. coli and B. japonicum. | Constitutive promoter for expression in E. coli and B. japonicum. | ||
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| + | For the silver BioBrick criteria, we decided to BioBrick and characterize the promoter from the plasmid pRJPaph-bjGFP (https://www.addgene.org/67032/). This promoter was used to express our pesticide resistance pathways and GFP in B.japonicum for our project this year. | ||
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<strong>The graph below indicates the results from this characterization experiment:</strong><br> | <strong>The graph below indicates the results from this characterization experiment:</strong><br> | ||
| − | [[File:T--waterloo--silvergraph.jpg|300px|thumb|left|Figure 1: OD-corrected GFP fluorescence | + | [[File:T--waterloo--silvergraph.jpg|300px|thumb|left|Figure 1: OD-corrected GFP fluorescence for characterization of (Excitation: 485, Emmission: 525)]] |
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| − | (Excitation: 485, Emmission: 525) | + | |
Revision as of 00:27, 22 October 2019
Paph promoter
Constitutive promoter for expression in E. coli and B. japonicum.
For the silver BioBrick criteria, we decided to BioBrick and characterize the promoter from the plasmid pRJPaph-bjGFP (https://www.addgene.org/67032/). This promoter was used to express our pesticide resistance pathways and GFP in B.japonicum for our project this year.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Characterization
(Waterloo iGEM 2019)
To characterize our promoter we cloned it upstream of the RBS in BBa_E0240 (RBS-GFP-Term) and compared it's GFP expression with the already well characterized BBa_J23101 promoter in BBa_I20260. To limit the impact of other variables we insured that BBa_I20260 and BBa_E0240 shared the same backbone, RBS and terminator. In addition to characterizing this promoter with respect toBBa_I20260 we also confirmed proper function of this promoter in the pRJPaph-bjGFP plasmid which was used for expression in Bradyrhizobium for our 2019 project.
Note:
empty DH5alpha was used as a negative control for GFP expression
1. Cell were grown overnight in LB media at 37 degrees
2. The following morning GFP expression was quantified using the Synergy 4 BioTek plate reader.
The graph below indicates the results from this characterization experiment:
