Difference between revisions of "Part:BBa K1789004"

 
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<partinfo>BBa_K1789004 short</partinfo>
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<partinfo>BBa_K1789009 short</partinfo>
  
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This part is the Carboxyl Half of GFP
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This is a engineered TAL effector that can bind with the DNA sequence 5'-GATAAACACCTTTC-3'
  
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<!-- Add more about the biology of this part here
 
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===Usage and Biology===
 
  
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==Usage and Biology==
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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.
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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].
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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]
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==Sequence and Features==
 
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<span class='h3bb'>Sequence and Features</span>
 
<span class='h3bb'>Sequence and Features</span>
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<partinfo>BBa_K1789004 SequenceAndFeatures</partinfo>
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<partinfo>BBa_K1789009 SequenceAndFeatures</partinfo>
  
  
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<!-- Uncomment this to enable Functional Parameter display
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===Functional Parameters===
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==Experimental Validation==
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<partinfo>BBa_K1789004 parameters</partinfo>
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After standardization, we used PCR to proved it.
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The forward primer sequence is 5’-CGGAATTCGCGGCCGCTTCTAGAAAGAATGGAATCC-3’.  The reverse primer sequence is 5’-GGACTAGTTTATTATTTGTATAGTTC -3’.
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[[File:GFP2_PCR|500px|]]
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As can be seen from the picture, it's length is right.
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we also sent it to sequencing, from the reporter we can proved that it's right.
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===References===
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[ 1] Drewes G, Bouwmeester T.Global approaches to protein – protein interaction[ J ]. Curr Opin Cell Biol, 2003, 15(2): 199-205.
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[ 2] Collura V, Boissy G. From protein -protein complexes to interactomics [ J ].Subcell Biochem, 2007, 43: 135-83.
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[ 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.

Revision as of 09:13, 18 September 2015

TALE2

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’.

File:GFP2 PCR

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.