Composite

Part:BBa_K2909014

Designed by: Laurent CHEN   Group: iGEM19_Sorbonne_U_Paris   (2019-08-28)

DGAT-1-2-HiBiT_LPAAT-A-HiBiT_HygroR E. guineensis

Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal EcoRI site found at 5568
    Illegal PstI site found at 2349
    Illegal PstI site found at 4307
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal EcoRI site found at 5568
    Illegal NheI site found at 5832
    Illegal PstI site found at 2349
    Illegal PstI site found at 4307
    Illegal NotI site found at 217
    Illegal NotI site found at 3291
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal EcoRI site found at 5568
    Illegal BglII site found at 2257
    Illegal BamHI site found at 4149
    Illegal XhoI site found at 758
    Illegal XhoI site found at 980
    Illegal XhoI site found at 3832
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal EcoRI site found at 5568
    Illegal PstI site found at 2349
    Illegal PstI site found at 4307
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal EcoRI site found at 5568
    Illegal PstI site found at 2349
    Illegal PstI site found at 4307
    Illegal NgoMIV site found at 4862
    Illegal NgoMIV site found at 6533
    Illegal NgoMIV site found at 6715
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 2550
    Illegal SapI.rc site found at 5039


Introduction

1- Biological background

This part is a composed of two CDS coding for the E. guineensis DGAT-1-2 (BBa_K2909003) and LPAAT-A (BBa_K2909002) enzymes, both tagged with a C-terminal HiBiT tag (BBa_K2909001).
Both enzymes are under the control of an ammonium-controlled Chlamydomonas reinhardtii promoter called NIT1.
This part also contains a Hygromycin resistance cassette called AphVII to select the recombinant clones.

This part has been designed to enhance the overall production of triglycerides in Chlamydomonas reinhardtii. The C-terminal HiBiT tag allows a quick determination of the enzyme expression.

This part is a linear DNA that is the result of the digestion of the level M plasmid by the type IIS BsaI restriction enzyme.
It is this linear DNA that is integrated into the genomic DNA of Chlamydomonas reinhardtii during the transformation.

2- Usage in iGEM projects

Bio(oil)gical Factory (iGEM Sorbonne Université 2019)

Characterization

References

  1. Schwinn, M. K. et al. CRISPR-Mediated Tagging of Endogenous Proteins with a Luminescent Peptide. ACS Chem. Biol. 13, 467–474 (2018).
  2. Crozet, P. et al. Birth of a Photosynthetic Chassis: A MoClo Toolkit Enabling Synthetic Biology in the Microalga Chlamydomonas reinhardtii. ACS Synth. Biol. 7, 2074–2086 (2018).
  3. 1.Weber, E., Engler, C., Gruetzner, R., Werner, S. & Marillonnet, S. A Modular Cloning System for Standardized Assembly of Multigene Constructs. PLoS ONE 6, e16765 (2011).
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Categories
Parameters
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