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

(→‎Gibson Assembly Design)
(→‎References and acknowledgements)
 
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==References and acknowledgements==
 
==References and acknowledgements==
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Thanks to the '''Mark Howarth laboratory at Oxford university Dept. of Biochemistry''' (link) for providing us with the streptavidin cloning plasmid.
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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.
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Thanks also to the '''iGEM team of Edinburgh 2011''' that designed the Biobrick BBa_K523013.
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<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].