Regulatory

Part:BBa_K2656007

Designed by: Adrian Requena Gutierrez, Carolina Ropero   Group: iGEM18_Valencia_UPV   (2018-09-16)

Constitutive promoter J23101

domestication.
Figure 1. DNA basic parts domestication. First construction refers to promoter domestication into the Golden Gate grammar.

Part BBa_K2656007 is the constitutive strong promoter BBa_J23101 adapted into the Golden Gate grammar. Thus, it is compatible with both Biobrick and Golden Braid assembly methods.

Following the Golden Gate assembly protocol , it can be combined with other GB compatible parts from our Valencia UPV IGEM 2018 Printeria Collection to assemble transcriptional units in a BsaI Type IIS one-pot reaction.

Characterization of the this part was performed with the transcriptional unit BBa_K2656107, which was used in a comparative promoters expression experiment with composite parts BBa_K2656105 and BBa_K2656107. They all were assembled in a Golden Braid alpha1 plasmid using the same RBS, CDS and terminator:

By using this experimental protocol, we have obtained the parameters to valide our constitutive model and so rationale choose its optimized values based on each promoter tested.

promoter experiment.
Figure 2. Promoter expression experiment with BBa_K2656004, K2656005 and BBa_K2656007 basic pieces


Table 1. Optimized parameters for the BBa_K2656007 promoter.
Parameter Value
Constitutive transcription rate CR CR = 486.25 min-1
Dilution rate μ μ = 0.0058 min-1

We have also calculated the relative force between the different promoters, taking BBa_K2656005 promoter as a reference. Likewise, the promoter ratio (p parameter ratio) of the different promoters referring to the stronger one has been calculated.

Table 2. BBa_K2656007 relative strength and p ratio.
Parameter Value
Relative strength 0.9367
p parameter ratio 0.9013


Sequence and Features

Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 11
    Illegal NheI site found at 34
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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Categories
Parameters
None