Part:BBa_K1433020

P-A-T-T-B-C-T-T-D-B0034-GFP-T

Lambda red is a Lambda-phage-derived recombination system which contains three proteins: Exo, Beta and Gam. It can recombine dsDNA/ssDNA into different kinds of DNA molecules such as chromosome, bacteria artificial chromosome (BAC) or even multicopy plasmids, as long as each side of the donor dsDNA/ssDNA is flanked by 36-50bp homologous arms. The homologous arms also determine the accurate sites of recombination, which ensures that the donor fragment can be inserted into any sites by choosing and adding homologous arms using PCR.

This part is designed to test lambda red recombination.

Composition:

1. Promoter: BBa_J23110, a middle-ground Bacterial constitutive promoter.
2. Homologous arms: homologous arms A, B, C and D.
3. Terminator: BBa_B0015, a strong double terminator.
4. RBS: BBa_B0034, a strong RBS.
5. GFP: BBa_E0040, green fluorescent protein derived from jellyfish Aequeora Victoria wild-type GFP.

In this circuit, four terminators suppress the expression of GFP, until recombination occurs twice between homologous arms A&B and C&D respectively. The inserted sequence replaces these four terminators and enables the expression of the downstream GFP. The green color of the strain can be easily observed.

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
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1393