Device

Part:BBa_K2017010

Designed by: Monica Victoria Gutierrez Salazar   Group: iGEM16_Valencia_UPV   (2016-10-04)
Revision as of 14:55, 4 October 2016 by Registry (Talk | contribs)

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35s + Ga20ox consense + RSIAT-TEV-Luciferase + Tnos

Modul of the gRNA testing system created by Valencia UPV 2016 team. This system aims to test the functionality and efficiency of a chosen gRNA to target a particular gene. This system is modular, as the gene inserted can be chosen by the user, only taking into account the required overhangs (5'-AATG-3' and 5'-TTCG-3').

It includes a promoter 35s (BBa_K1537015), 20 nucleotides of the Ga20ox consensus gene of Oryza sativa (rice), the reporter luciferase with RSIAT+TEV linker and the terminator Tnos.

Luciferase is out of its reading frame due to a nucleotide added upstream to the linker, and the initial ATG has been removed. This means that when the luciferase is translated, it will not be functional. To make it functional, it is necessary to make indels in the Ga20ox fragment that put the luciferase in the correct reading frame. These indels can be made using the CRISPR/Cas9 system, which is the aim of this device.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 871
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 66
    Illegal BsaI site found at 2577
    Illegal BsaI.rc site found at 2864
    Illegal SapI.rc site found at 1723


[edit]
Categories
//cds/reporter
//chassis/eukaryote/nbenthamiana
//function/crispr
Parameters
None