Difference between revisions of "Part:BBa K649206"

Latest revision as of 05:25, 29 October 2011

lox66

To characterize this part, we constructed the generator part BBa_K649202.

The lox sequences, lox71 and lox66, have 5 bp on the 5 and 3 ends changed, respectively. DNA segment flanked by lox71 and lox66 marks the point which the enzyme Cre will excise. This Part is expected to express GFP when the lox sites are excised and RFP when they are not. The Cre-mediated recombination of this BioBrick had been studied and proved to be working.

Effect of Cre-meditated recombination at lox71/66 cassette.
The leftmost is a negative control which don'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
COMPATIBLE WITH RFC[21]
23
COMPATIBLE WITH RFC[23]
25
COMPATIBLE WITH RFC[25]
1000
COMPATIBLE WITH RFC[1000]

@@ Line 4: / Line 4: @@
 To characterize this part, we constructed the generator part [https://parts.igem.org/Part:BBa_K649202 BBa_K649202].
-The lox sequences, lox71 and lox66, have 5 bp on the 5 and 3 ends changed, respectively. DNA segment flanked by lox71 and lox66 marks the point which the enzyme Cre will excise. This Part is expected to express GFP when the lox sites are excised and RFP when they are not. The Cre-mediated recombination of this BioBrick had been studied and proved to be working.
+The ''lox'' sequences, ''lox71'' and ''lox66'', have 5 bp on the 5 and 3 ends changed, respectively. DNA segment flanked by ''lox71'' and ''lox66'' marks the point which the enzyme Cre will excise. This Part is expected to express GFP when the ''lox'' sites are excised and RFP when they are not. The Cre-mediated recombination of this BioBrick had been studied and proved to be working.
-[[Image:111001DK_1hr_D7166_111003.png|thumb|center|500px|Effect of Cre-meditated recombination at lox71/66 cassette.<br />The leftmost is a negative control which don'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.
+[[Image:111001DK_1hr_D7166_111003.png|thumb|center|500px|Effect of Cre-meditated recombination at ''lox71/66'' cassette.<br />The leftmost is a negative control which don'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.
 ]]
-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.
+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.