Generator

Part:BBa_K814001

Designed by: Nolan Kennedy, Michelle Goettge, Grayson Wawrzyn   Group: iGEM12_Minnesota   (2012-10-03)

ATP-grasp (ATPG) generator

Our research has focused on two novel biosynthetic pathways found in two distinct algal species. A pathway ending in the production of two UV-protective compounds, shinorine and mycosporine-glycine, was cloned from Anabaena varibalis. ATP-grasp (ATPG) catalyzes the third step in this pathway, 4-deoxygadusol into mycosporine-glycine.

This part includes a modified constitutive lac promoter (lacP'), and RBS and the open reading frame of DHQS. This part can be used to express DHQS in E. coli. Dhqs.png

Figure 1. PCR screen of ATP-grasp confirming successful cloning of this ORF into the pUCBB plasmid.

Balskus and Walsh, 2010. [1]

BBa_K3634002

Codon optimised adenosine triphosphate (ATP)-grasp (ATPG) CDS for use in E.coli as part of the shinorine synthesis gene cluster. By using the IDT codon optimisation tool, St Andrews iGEM 2020 have optimised the GC content of the sequence taken from A.variabilis to improve production efficiency of the final product shinorine. We have further made the part biobrick assembly standard RFC[10] & RFC[1000] compatible by removing EcoR1 and PstI restriction sites introduced by this optimisation step. The part should be used alongside the additional optimised parts (BBa_K3634000, BBa_K3634001 and BBa_K3634003) responsible for the ultimate conversion of the substrate sedoheptulose 7-phosphate to the final product shinorine.

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 944
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 944
    Illegal NotI site found at 1517
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 944
    Illegal BglII site found at 134
    Illegal XhoI site found at 1525
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 944
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 944
  • 1000
    COMPATIBLE WITH RFC[1000]


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Categories
Parameters
None