Part:BBa_K2909012

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)

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).