Composite

Part:BBa_K2909012

Designed by: Laurent CHEN   Group: iGEM19_Sorbonne_U_Paris   (2019-08-28)
Revision as of 08:24, 29 August 2019 by Laurent CHEN (Talk | contribs) (→‎Introduction)

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DGAT-1-2-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 PstI site found at 2349
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal NheI site found at 3343
    Illegal PstI site found at 2349
    Illegal NotI site found at 217
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal BglII site found at 2257
    Illegal XhoI site found at 758
    Illegal XhoI site found at 980
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal PstI site found at 2349
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3079
    Illegal PstI site found at 2349
    Illegal NgoMIV site found at 4044
    Illegal NgoMIV site found at 4226
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 2550


Introduction

1- Biological background

This part is a CDS coding for the E. guineensis DGAT-1-2 enzyme tagged with a C-terminal HiBiT tag under the control of an ammonium-controlled Chlamydomonas reinhardtii promoter called NIT1. It also contains a Hygromycin resistance cassette called AphVII to select the recombinant clones.

This part has been designed to characterize the expression of the E. guineensis DGAT-1-2 enzyme that we adapted for C. reinhardtii (BBa_K2909003) using the C-terminal HiBiT tag (BBa_K2909001).

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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