Difference between revisions of "Part:BBa K1321334:Design"

 
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<partinfo>BBa_K1321334 short</partinfo>
 
<partinfo>BBa_K1321334 short</partinfo>
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===Design Notes===
 
===Design Notes===
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The Ribosomal binding site of AcsAB is already included within the sequence. Because we are transferring this element from Gluconacetobacter into simpler organisms, such as E.coli, the native RBS was replaced with a new RBS, that would guarantee strong translation whilst maintaining the native stoichiometry for the final operon system to operate successfully.  
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The AcsAB coding sequence was extracted from NCBI (refer to Source). Some native regulation was removed by codon-optimisation for expression in <em>E.coli</em>. The native ribosomal binding sites (RBS) were predicted to be weak by translation rate calculators (Sallis Lab RBS calculator and Postech UTR designer), therefore they were replaced by the B0034 strong RBS. Furthermore, the six base pairs downstream and upstream from this RBS were edited with the Sallis Lab RBS calculator so as to tune RBS strength and try to preserve the native stoichiometry.  
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===Source===
 
===Source===
  
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The AcsAB coding sequence was extracted from NCBI, codon optimised and edited accordingly for further gene synthesis and cloning into pSB1C3.  
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<li>http://www.ncbi.nlm.nih.gov/nuccore/X54676.1
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<li>https://salis.psu.edu/software/
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<li>http://sbi.postech.ac.kr/utr_designer
  
 
===References===
 
===References===
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Lee, K.Y; Guldum, G.; Mantalaris, A.; Bismarck, A.; (2014) “More than meets the eye in Bacterial Cellulose: Biosynthesis, Bioprocessing and Applications in Advanced Fiber Composites” Available on: http://onlinelibrary.wiley.com/store/10.1002/mabi.201300298/asset/mabi201300298.pdf?v=1&t=i1i6d96u&s=ca0d79f542dfb621dcecf8e90feefb7dd06d031c

Latest revision as of 18:48, 20 October 2014

pSB-AcsAB


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1831
    Illegal BglII site found at 2545
    Illegal BglII site found at 4468
    Illegal BamHI site found at 841
    Illegal BamHI site found at 3835
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 1817
    Illegal AgeI site found at 1894
    Illegal AgeI site found at 2450
    Illegal AgeI site found at 3251
    Illegal AgeI site found at 3343
    Illegal AgeI site found at 3449
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

The AcsAB coding sequence was extracted from NCBI (refer to Source). Some native regulation was removed by codon-optimisation for expression in E.coli. The native ribosomal binding sites (RBS) were predicted to be weak by translation rate calculators (Sallis Lab RBS calculator and Postech UTR designer), therefore they were replaced by the B0034 strong RBS. Furthermore, the six base pairs downstream and upstream from this RBS were edited with the Sallis Lab RBS calculator so as to tune RBS strength and try to preserve the native stoichiometry.

Source

  • http://www.ncbi.nlm.nih.gov/nuccore/X54676.1
  • https://salis.psu.edu/software/
  • http://sbi.postech.ac.kr/utr_designer

    References

    Lee, K.Y; Guldum, G.; Mantalaris, A.; Bismarck, A.; (2014) “More than meets the eye in Bacterial Cellulose: Biosynthesis, Bioprocessing and Applications in Advanced Fiber Composites” Available on: http://onlinelibrary.wiley.com/store/10.1002/mabi.201300298/asset/mabi201300298.pdf?v=1&t=i1i6d96u&s=ca0d79f542dfb621dcecf8e90feefb7dd06d031c