Part:BBa_K5023014

PAR_Ble_SP7_FAST-PETase_Linker_MHETase_SP20_Ter

This composite part have: PAR promoter, the bleomycin/zeocin resistance sequence, signal peptide, the PHL7 enzyme and the terminator. Designed to express and secrete PHL7 in Chlamydomonas reinhardtii, utilizing pJP32 backbone. This circuit was originally designed by our advisor João Molino.

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 698
  Illegal PstI site found at 1572
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 249
  Illegal NheI site found at 1211
  Illegal PstI site found at 698
  Illegal PstI site found at 1572
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal XhoI site found at 2043
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 698
  Illegal PstI site found at 1572
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 698
  Illegal PstI site found at 1572
  Illegal NgoMIV site found at 1079
  Illegal NgoMIV site found at 1140
  Illegal NgoMIV site found at 1357
  Illegal NgoMIV site found at 1458
  Illegal NgoMIV site found at 2370
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1808
  

Introduction to the Mhetygua Project

The Mhetygua project, initiated for the IGEM Global 2023 edition, represents a pioneering endeavor to combat the pressing issue of microplastic pollution. Spearheaded by a dedicated group of students, the project titled "MHETYGUÁ: algae as devices for the degradation of aquatic pollutants" seeks to harness the potential of algae, specifically Chlamydomonas reinhardtii, as a bio-tool for microplastic degradation. Beyond the scientific pursuit, the project embodies a broader vision of fostering environmental awareness and education, aiming to catalyze a shift towards sustainable practices and a reduced plastic footprint in the Latin American region.

Commentary on the Genetic Circuit Construction: Circuit Number 4

The genetic circuit, labeled as Circuit Number 4, is a unique design aimed at achieving the expression and secretion of the enzyme PHL7 in the microalgae Chlamydomonas reinhardtii. This circuit was originally conceptualized by our esteemed advisor, João Molino. The construction of this circuit involves the following key components:

1. PAR Promoter: The PAR promoter initiates the transcription process. Recognized for its light-inducible properties, it allows for the modulation of gene expression based on light conditions, making it particularly effective in Chlamydomonas reinhardtii.
2. Bleomycin/Zeocin Resistance Sequence: This sequence provides resistance against the antibiotics bleomycin and zeocin. Its inclusion ensures that only algal cells with the integrated genetic circuit can thrive in the presence of these antibiotics, facilitating the selection of desired traits.
3. Signal Peptide: Essential for the secretion of the synthesized enzyme, the signal peptide guides the PHL7 protein towards the secretory pathway, ensuring its release into the surrounding environment.
4. PHL7 Enzyme: PHL7 is the focal enzyme of this circuit. Once expressed and secreted, it is expected to perform its designated biochemical functions, contributing to the overarching goals of the Mhetygua project.
5. Terminator: The terminator sequence marks the end of the transcription process, ensuring that the genetic expression is both stable and efficient.

The entire circuit is housed within the pJP32 backbone. This vector is chosen for its compatibility with Chlamydomonas reinhardtii, ensuring that the circuit is successfully integrated and expressed within the algal cells.