Difference between revisions of "Part:BBa K2995000"
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To characterize our promoter we cloned it upstream of the RBS in <partinfo>BBa_E0240</partinfo> (RBS-GFP-Term) and compared it's GFP expression with the already well characterized <partinfo>BBa_J23101</partinfo> promoter in <partinfo>BBa_I20260</partinfo>. To limit the impact of other variables we insured that <partinfo>BBa_I20260</partinfo> and <partinfo>BBa_E0240</partinfo> shared the same backbone, RBS and terminator. In addition to characterizing this promoter with respect to<partinfo>BBa_I20260</partinfo> we also confirmed proper function of this promoter in the pRJPaph-bjGFP plasmid which was used for expression in Bradyrhizobium for our 2019 project.   
 
To characterize our promoter we cloned it upstream of the RBS in <partinfo>BBa_E0240</partinfo> (RBS-GFP-Term) and compared it's GFP expression with the already well characterized <partinfo>BBa_J23101</partinfo> promoter in <partinfo>BBa_I20260</partinfo>. To limit the impact of other variables we insured that <partinfo>BBa_I20260</partinfo> and <partinfo>BBa_E0240</partinfo> shared the same backbone, RBS and terminator. In addition to characterizing this promoter with respect to<partinfo>BBa_I20260</partinfo> we also confirmed proper function of this promoter in the pRJPaph-bjGFP plasmid which was used for expression in Bradyrhizobium for our 2019 project.   
  
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==Note:== empty DH5alpha was used as a negative control for GFP expression
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<strong>Note:</strong><br> empty DH5alpha was used as a negative control for GFP expression
  
 
1. Cell were grown overnight in LB media at 37 degrees
 
1. Cell were grown overnight in LB media at 37 degrees

Revision as of 23:22, 21 October 2019


Paph promoter

Constitutive promoter for expression in E. coli and B. japonicum.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE 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:


(Excitation: 485, Emmission: 525)