Part:BBa_K3866000

araC-ParaBAD:RBS GB compatible with A1-B2

Figure 1. The level 0 module : pupd2- araC:araBAD (illustration from SnapGene)

Usage and Biology

The PBAD promoter, derived from the arabinose operon andis an inducible promoter for arabinose. The use of an inducible promoter provides a most reversible and flexible gene circuit and at the same time exhibits a higher efficiency and lower side effects as cell death.(Kallunki, Barisic, Jäättelä and Liu, 2019). The araC gene, is a regulatory gene and is located upstream of the L-arabinose operon, encodes a positive regulatory protein AraC, required for L-arabinose utilization in Escherichia coli. The promoter is highly inhibited from Glucose. The araC gene has a constitutive promoter.

Design Notes

The coding sequence was domesticated . We removed BsmBI ,BsaI , BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is present in pUPD2 BBa_K3505007 and has overhangs compatible for GoldenBraid cloning. The CDS has position A1-B2

Figure 2.The overhangs of this part in the GoldenBraid Grammar.

Verification of Cloning

Fig.3:(U=Uncut C=Cut) Restriction Digestion of araC:ParaBAD with EcoRI+ BamHI. Positive clones 1, 2 Expected bands 2160, 1184

Experimental Use and Experience

This part showed fuctionality at the following parts. BBa_K3866009 BBa_K3866010 BBa_K3866026 BBa_K3866027 BBa_K3866028

Source

From the iGEM Distribution Kit 2019.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 1148
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 983
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal SapI site found at 965
  

References

• Kallunki, Barisic, Jäättelä and Liu, 2019. How to Choose the Right Inducible Gene ExpressionSystem for Mammalian Studies?Cells, 8(8), p.796.