Difference between revisions of "Part:BBa K1111012:Design"

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==Gibson Assembly Design ==
 
==Gibson Assembly Design ==
−
Insert: we amplified the streptavidin coding sequence received in a plasmid from Mark Howarth laboratory at Oxford university Dept. of Biochemistry.  
+
''Insert:'' we amplified the streptavidin coding sequence received in a plasmid from Mark Howarth laboratory at Oxford university Dept. of Biochemistry.  
−
Backbone: Starting from the biobrick BBa_K523013 (INP fused with YFP), we amplified the whole sequence except the EYFP and added gibson overhang complementary to streptavidin ends.
+
<br>''Backbone:'' Starting from the biobrick BBa_K523013 (INP fused with YFP), we amplified the whole sequence except the EYFP and added gibson overhang complementary to streptavidin ends.
  
 
==Primers==
 
==Primers==
−
PCR of streptavidin alive :
+
Streptavidin Alive PCR :
−
5' ATGGCTGAAGCTGGTATCACC 3'
+
<br>5' <font color = #0000FF>ATGGCTGAAGCTGGTATCACC</font> 3'
−
5' TTAGGAAGCAGCGGACGGTTTAAC 3'
+
<br>5' <font color = #0000FF>TTAGGAAGCAGCGGACGGTTTAAC</font> 3'
  
−
Overlapping PCR of BBa_K523013 :  
+
BBa_K523013 PCR :  
−
Fw: 5' <font color = bleue> accaaagttaaaccgtccgctgcttcct</font><font color= red>aacatatcataacggagtgatcgcaatg</font> 3'
+
<br>Fw: 5' <font color = #0000FF>ACCAAAGTTAAACCGTCCGCTGCTTCT</font><font color= red>AACATATCATAACGGAGTGATCGCAATG</font> 3'
−
Rev: 5' ccaggtgccggtgataccagcttcagcCATagatcccgccacgctgct 3'
+
<br>Rev: 5' <font color = #0000FF>CCAGGTGCCGGTGATACCAGCTTCAGCCAT</font><font color= red>AGATCCCGCCACGCTGCT</font> 3'
 +
 
 +
==Source==
 +
Can be expressed in Escherichia Coli.
  
 
==References and acknowledgements==
 
==References and acknowledgements==
−
Thanks to the '''Mark Howarth laboratory at Oxford university Dept. of Biochemistry''' (link) for providing us with the streptavidin cloning plasmid.
+
Thanks to the '''Mark Howarth laboratory at Oxford university Dept. of Biochemistry''' [http://users.ox.ac.uk/~bioc0756/MyWebs/activesite/ReagentDistribution.htm] for providing us with the streptavidin cloning plasmid.
−
Thanks also to the '''iGEM team of Edinburgh 2011''' that designed the Biobrick BBa_K523013.
+
<br>Thanks to the '''iGEM team of Edinburgh 2011''' that designed the Biobrick BBa_K523013 [https://parts.igem.org/Part:BBa_K523013].

Latest revision as of 21:41, 2 October 2013

Ice Nucleation Protein fused to Streptavidin Alive


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal XbaI site found at 1727
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1649
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal XbaI site found at 1727
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal XbaI site found at 1727
    Illegal NgoMIV site found at 1036
    Illegal AgeI site found at 1691
    Illegal AgeI site found at 1742
  • 1000
    COMPATIBLE WITH RFC[1000]

Gibson Assembly Design

Insert: we amplified the streptavidin coding sequence received in a plasmid from Mark Howarth laboratory at Oxford university Dept. of Biochemistry.
Backbone: Starting from the biobrick BBa_K523013 (INP fused with YFP), we amplified the whole sequence except the EYFP and added gibson overhang complementary to streptavidin ends.

Primers

Streptavidin Alive PCR :
5' ATGGCTGAAGCTGGTATCACC 3'
5' TTAGGAAGCAGCGGACGGTTTAAC 3'

BBa_K523013 PCR :
Fw: 5' ACCAAAGTTAAACCGTCCGCTGCTTCTAACATATCATAACGGAGTGATCGCAATG 3'
Rev: 5' CCAGGTGCCGGTGATACCAGCTTCAGCCATAGATCCCGCCACGCTGCT 3'

Source

Can be expressed in Escherichia Coli.

References and acknowledgements

Thanks to the Mark Howarth laboratory at Oxford university Dept. of Biochemistry [http://users.ox.ac.uk/~bioc0756/MyWebs/activesite/ReagentDistribution.htm] for providing us with the streptavidin cloning plasmid.
Thanks to the iGEM team of Edinburgh 2011 that designed the Biobrick BBa_K523013 [1].