Part:BBa_K137054

Constitutive folKE expression construct

This is the folate synthesis gene folKE, one of five genes (folB, folKE, folP, folC, folA) involved in the folate synthesis operon. In this construct, folKE has been inserted behind the strong promoter J23100, the strong RBS B0034, and in front of the double terminator B0015. The vector is the high copy plasmid pSB1A2.

The purpose of this construct is to test the effects of FolKE (a bifunctional GTP cyclohydrolase I (FolE) + 2-amino-4-hydroxy-6- hydroxymethyldihydropteridine pyrophosphokinase (FolK)) overexpression on total folate production in Escherichia coli, with the goal of increasing total folate production.

Usage and Biology

Using the Caltech iGEM [http://2008.igem.org/Team:Caltech/Protocols/Folate_assay microbiological folate assay], K137054 in pSB1A2 in DH10B produced the following results:

When supplemented with PABA, K137054 produced more growth in the microbiological assay, indicating a higher production of folate. Cultures without K137054 or without PABA produced lower growth (indicating lower levels of folate production).

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]