Composite

Part:BBa_K2909013:Design

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

HiBiT-DGAT-1-2_HiBiT-LPAAT-A_HygroR E. guineensis


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3104
    Illegal EcoRI site found at 5618
    Illegal PstI site found at 2389
    Illegal PstI site found at 4372
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3104
    Illegal EcoRI site found at 5618
    Illegal NheI site found at 5882
    Illegal PstI site found at 2389
    Illegal PstI site found at 4372
    Illegal NotI site found at 217
    Illegal NotI site found at 3316
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3104
    Illegal EcoRI site found at 5618
    Illegal BglII site found at 2297
    Illegal BamHI site found at 2537
    Illegal BamHI site found at 4214
    Illegal BamHI site found at 5051
    Illegal XhoI site found at 758
    Illegal XhoI site found at 1020
    Illegal XhoI site found at 3857
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3104
    Illegal EcoRI site found at 5618
    Illegal PstI site found at 2389
    Illegal PstI site found at 4372
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal EcoRI site found at 5
    Illegal EcoRI site found at 3104
    Illegal EcoRI site found at 5618
    Illegal PstI site found at 2389
    Illegal PstI site found at 4372
    Illegal NgoMIV site found at 4927
    Illegal NgoMIV site found at 6583
    Illegal NgoMIV site found at 6765
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 2575
    Illegal SapI.rc site found at 5089


Design Notes

Source

The LPAAT-A enzyme (BBa_K2909002) comes from the predicted sequence from the E. guineensis genome sequencing.
NCBI Reference Sequence: XM_010908457.2
https://www.ncbi.nlm.nih.gov/nuccore/1130625520

The DGAT-1-2 enzyme (BBa_K2909003) comes from the predicted sequence from the E. guineensis genome sequencing.
NCBI Reference Sequence: XM_010933834.1
https://www.ncbi.nlm.nih.gov/nuccore/XM_010933834.1

The N-terminal HiBiT tag (BBa_K2909000) comes from Promega (Schwinn et al. 2018).

All the other parts comes from the Chlamydomonas reinhardtii MoClo Kit (Crozet et al. 2018).

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).