Composite

Part:BBa_K2353002:Design

Designed by: Julia Leveille, Lauren Hong, Emily Gibson, Gaurav Byagathvalli, Katie Barr, Alyssa Franklin, Christina Lee, Ellie Kim, Kevin Li, Megan Hong, Natalie Shih, Nithik Balachandran, Noora Chandasir, David S   Group: iGEM17_Lambert_GA   (2017-10-17)
Revision as of 16:10, 1 November 2017 by JLeveille (Talk | contribs) (→‎References)


tsPurpleLAA


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


Design Notes

tsPurpleLAA is composed of a ribosomal binding site, tsPurple (a purple chromoprotein which functions as a reporter), and LAA (a fast degradation tag). tsPurpleLAA is intended to be assembled with pLambdaR-LacI (which is the promoter and regulatory system) and r0011ClpXPCI (which is the non-lysosomal protease). The terminator is located at the beginning of pLac-ClpXP-CI. When fully assembled, p-lambda-r LacI transcribes tsPurpleLAA and LacI represses pLac, preventing transcription of ClpXP-CI. Upon induction with IPTG, LacI binds to IPTG which prevents repression of pLac. Therefore, ClpXP-CI is transcribed; ClpXP should recognize the LAA deg tag and degrade the tsPurple protein. Since LAA is a fast degradation tag, so tsPurpleLAA should have the highest levels of degradation seen. The ensuing levels of expression could then be compared to that of BBa_K2353000 (tsPurple without a deg tag) to see the relative level of degradation experienced.

IMPORTANT NOTE: The promoter and terminator are both part of different parts, which means tsPurpleLAA is not expressed unless it is assembled with pLambdaR-LacI (BBa_K1911000). The protease complex of ClpX and ClpP is from part (BBa_K1911001).

Source

This part is composed from sequences found in the parts registry.


References

Eurofins Genomics. (n.d.) Retrieved November 01, 2017, from https://www.eurofins.com Chromogenic Protein Paintbox™ - E. coli – pMOTHER vectors. (n.d.) Retrieved November 01, 2017, from https://www.atum.bio/eCommerce/catalog/datasheet/526 Part:BBa_K1033906. (n.d.). Retrieved November 01, 2017, from https://parts.igem.org/wiki/index.php?title=Part%3ABBa_K1033906 Part:BBa_M0050. (n.d.) Retrieved November 01, 2017, from https://parts.igem.org/Part:BBa_M0050