Difference between revisions of "Part:BBa K243010:Design"
Line 6: Line 6:
  
 
===Design Notes===
 
===Design Notes===
−
The cloning steps were planed theoretically before we started the work in the wet lab.
+
The cloning steps were planned theoretically before we started the work in the wet lab.
−
The combiantion of HisTag linked with a FluA Tag connect by the Split Linker approved as a good way to express the [https://parts.igem.org/wiki/index.php?title=Part:BBa_K243001 inactive protein domain] of our universal endonuclease.<br><br>
+
The combination of HisTag linked with a FluA tag connected by the Split Linker approved as a good way to express the [https://parts.igem.org/wiki/index.php?title=Part:BBa_K243001 inactive protein domain] of our universal endonuclease.<br><br>
 
[[Image:Freiburg09 Hisfluasplfoki.jpg|550x350px]]<br>
 
[[Image:Freiburg09 Hisfluasplfoki.jpg|550x350px]]<br>
 
<p class=MsoNormal><b>part of universal restriction enzyme.</b> Blue:
 
<p class=MsoNormal><b>part of universal restriction enzyme.</b> Blue:
 
DNA strand; Red: 16bp long Oligos, tag as indicated in the picture.<br> Ochre:
 
DNA strand; Red: 16bp long Oligos, tag as indicated in the picture.<br> Ochre:
 
Fluorescein A binding lipocalin;light Blue: inactive FokI cleavage domain;</p>   
 
Fluorescein A binding lipocalin;light Blue: inactive FokI cleavage domain;</p>   
−
We used the [https://parts.igem.org/Part:BBa_K157009 split linker], because it is an improved part of the team [http://2008.igem.org/Team:Freiburg Freiburg08] and it is a suitable linker for fusion proteins. The properties of the split linker are an good compromise between the stability and the distance for the connection between protein domain Fok_a and the anticaline tag.  
+
We used the [https://parts.igem.org/Part:BBa_K157009 split linker] because it is an improved part of the team [http://2008.igem.org/Team:Freiburg Freiburg08] and it is a suitable linker for fusion proteins. The properties of the split linker are a good compromise between the stability and the distance for the connection between protein domain Fok_a and the anticaline tag.  
 
The parts are fused according to [https://parts.igem.org/Assembly_standard_25 RFC 25].<br>
 
The parts are fused according to [https://parts.igem.org/Assembly_standard_25 RFC 25].<br>
 
[https://static.igem.org/mediawiki/parts/6/6a/HisFluASplitFoki.txt Commented GenBank file]
 
[https://static.igem.org/mediawiki/parts/6/6a/HisFluASplitFoki.txt Commented GenBank file]

Revision as of 20:57, 21 October 2009

His-FluA-Split Linker-Fok_i


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 272
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The cloning steps were planned theoretically before we started the work in the wet lab. The combination of HisTag linked with a FluA tag connected by the Split Linker approved as a good way to express the inactive protein domain of our universal endonuclease.

Freiburg09 Hisfluasplfoki.jpg

part of universal restriction enzyme. Blue: DNA strand; Red: 16bp long Oligos, tag as indicated in the picture.
Ochre: Fluorescein A binding lipocalin;light Blue: inactive FokI cleavage domain;

We used the split linker because it is an improved part of the team [http://2008.igem.org/Team:Freiburg Freiburg08] and it is a suitable linker for fusion proteins. The properties of the split linker are a good compromise between the stability and the distance for the connection between protein domain Fok_a and the anticaline tag. The parts are fused according to RFC 25.
Commented GenBank file

Source

Combined the parts by serial cloning steps.

References