Part:BBa_M50083:Design

Modified Yeast Serotonin Receptor

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 780
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 150
  
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

The overall goal was to express a human serotonin receptor in yeast (Saccharomyces cerevisiae). We sourced the serotonin receptor amino acid sequence from the human genome, and codon-optimized it to yeast. We added a HIS-tag to the N-terminus side of the gene as the extracellular side to enable testing for correct localization. Since we suspected the protein might be toxic to the yeast cells, we added an inducible promoter, rather than constituent, to control the level of expression. We used standard yeast Kozak and terminator sequences to allow for predictable transcription and translation.

Source

The serotonin receptor gene comes from the human genome. We took the amino acid sequence from http://www.uniprot.org/uniprot/P08908#sequences, and then simply codon optimized for yeast to generate the nucleotide sequence. The other parts are standard parts that can be found documented in the iGem Parts Registry.

References

Nakamura, Y., Ishii, J. and Kondo, A. (2015), Applications of yeast-based signaling sensor for characterization of antagonist and analysis of site-directed mutants of the human serotonin 1A receptor. Biotechnol. Bioeng., 112: 1906–1915. doi:10.1002/bit.25597

Part:BBa_J63006:Experience. Registry of Standard Biological Parts. https://parts.igem.org/Part:BBa_J63006:Experience. Online