Difference between revisions of "Part:BBa K649207"
 
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<partinfo>BBa_K649207 short</partinfo>
 
<partinfo>BBa_K649207 short</partinfo>
  
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PlacIQ-lox2272-gfp
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PlacIQ-''lox2272-gfp''
  
 
To characterize this part, we constructed the generator part [https://parts.igem.org/wiki/index.php?title=Part:BBa_K649201 BBa_K649201].
 
To characterize this part, we constructed the generator part [https://parts.igem.org/wiki/index.php?title=Part:BBa_K649201 BBa_K649201].
  
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lox2272 is a mutant of loxP which has mutations in the 8 bp spacer region. DNA segment flanked by two lox2272 marks the point which the enzyme Cre will excise. The Cre-mediated recombination of this BioBrick had been studied and proved to be working.
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''lox2272'' is a mutant of ''loxP'' which has mutations in the 8 bp spacer region. DNA segment flanked by two ''lox2272'' marks the point which the enzyme Cre will excise. The Cre-mediated recombination of this BioBrick had been studied and proved to be working.
  
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[[Image:111001DK_0.5_2272.png|thumb|center|500px|Cre-meditated recombination at lox2272 cassette.<br />The leftmost is a negative control which doesn't have Cre-expressing plasmid. <br />The center is an arabinose induced sample which has both Cre plasmid and BioBrick BBa_K649201. <br />The rightmost is a uninduced strain which has both plasmid like as the center.
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[[Image:111001DK_0.5_2272.png|thumb|center|500px|Cre-meditated recombination at ''lox2272'' cassette.<br />The leftmost is a negative control which doesn't have Cre-expressing plasmid. <br />The center is an arabinose induced sample which has both Cre plasmid and BioBrick BBa_K649201. <br />The rightmost is a uninduced strain which has both plasmid like as the center.
 
]]
 
]]
  
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In ''in vivo'' assay, arabinose induced strain which has Cre-expressing plasmid([[https://parts.igem.org/Part:BBa_I718008]]PBAD/araC-Cre, BBa_I718008) was expressing GFP, while negative control which doesn't have Cre plasmid was expressing Red florescence. It means that DNA recombination did happen by Cre recombinase. The uninduced sample was supposed to unexpress Cre recombinase, but because of the leaking of PBAD/araC promoter, the excision occured in almost every cell in medium.
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In ''in vivo'' assay, arabinose induced strain which has Cre-expressing plasmid([https://parts.igem.org/Part:BBa_I718008 PBAD/araC-Cre, BBa_I718008]) was expressing GFP, while negative control which doesn't have Cre plasmid was expressing Red florescence. It means that DNA recombination did happen by Cre recombinase. The uninduced sample was supposed to unexpress Cre recombinase, but because of the leaking of PBAD/araC promoter, the excision occured in almost every cell in medium.
  
 
For more information, see [http://2011.igem.org/Team:Tokyo_Tech/Projects/RPS-game/assay#8. our work in Tokyo_Tech 2011 wiki].
 
For more information, see [http://2011.igem.org/Team:Tokyo_Tech/Projects/RPS-game/assay#8. our work in Tokyo_Tech 2011 wiki].

Latest revision as of 05:33, 29 October 2011

PlacIQ-lox2272-gfp

PlacIQ-lox2272-gfp

To characterize this part, we constructed the generator part BBa_K649201.

lox2272 is a mutant of loxP which has mutations in the 8 bp spacer region. DNA segment flanked by two lox2272 marks the point which the enzyme Cre will excise. The Cre-mediated recombination of this BioBrick had been studied and proved to be working.

Cre-meditated recombination at lox2272 cassette.
The leftmost is a negative control which doesn't have Cre-expressing plasmid.
The center is an arabinose induced sample which has both Cre plasmid and BioBrick BBa_K649201.
The rightmost is a uninduced strain which has both plasmid like as the center.

In in vivo assay, arabinose induced strain which has Cre-expressing plasmid(PBAD/araC-Cre, BBa_I718008) was expressing GFP, while negative control which doesn't have Cre plasmid was expressing Red florescence. It means that DNA recombination did happen by Cre recombinase. The uninduced sample was supposed to unexpress Cre recombinase, but because of the leaking of PBAD/araC promoter, the excision occured in almost every cell in medium.

For more information, see [http://2011.igem.org/Team:Tokyo_Tech/Projects/RPS-game/assay#8. our work in Tokyo_Tech 2011 wiki].


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 107
    Illegal BamHI site found at 13
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 790