Part:BBa_K1789004

GFP2

This is a engineered TAL effector that can bind with the DNA sequence 5'-GATAAACACCTTTC-3'

Usage and Biology

Bimolecular fluorescence complementation (BiFC) means two non-fluorescent complementary fragments of the fluorescent protein can reassemble to form a fluorescent complex and restore fluorescence when they are fused to two proteins that interact with each other.

BiFC analysis has been used to study interactions among a wide range of proteins in many cell types. The study of interactions and post-translational modification of the protein makes people master the biological regulatory mechanism more. Interactions in protein are also highly valued, so there have been a number of related technologies having different characteristics and applications[1-2].

Such as GFP,after the sequence of certain sites in interchanges with the amino-terminal or carboxy-terminal sequence loop, it could still be able to fold correctly to form the structure of the chromophore and maintain fluorescence properties[3-5]

Sequence and Features

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 2083
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 1177
  Illegal BsaI.rc site found at 1381
  

Experimental Validation

After standardization, we used PCR to proved it.

The forward primer sequence is 5’-CGGAATTCGCGGCCGCTTCTAGAAAGAATGGAATCC-3’. The reverse primer sequence is 5’-GGACTAGTTTATTATTTGTATAGTTC -3’.

As can be seen from the picture, it's length is right.

we also sent it to sequencing, from the reporter we can proved that it's right.

References

[ 1] Drewes G, Bouwmeester T.Global approaches to protein – protein interaction[ J ]. Curr Opin Cell Biol, 2003, 15(2): 199-205.

[ 2] Collura V, Boissy G. From protein -protein complexes to interactomics [ J ].Subcell Biochem, 2007, 43: 135-83.

[ 3] M isteli T, Spector DL. Application of the green fluorescent protein in cell biology and biotechnology[ J ].Nat Biotechnol, 1997, 15( 10): 961 - 964.

[ 4] Baird G. S, Zacharias DA, Tsien RY. Circular permutation and receptor insertion within green fluorescent proteins[ J ]. Proc Natl Acad Sci USA , 1999, 96( 20): 11241-11246.

[ 5] Ghosh I, Hamilton AD, Regan L. Antiparallel leucinezipper –directed protein reassembly: application to the green fluorescent protein[ J ]. J Am Chem Soc, 2000, 122: 5658-5659.