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Coding

Part:BBa_K1323010

Designed by: Julia Manalil   Group: iGEM14_Waterloo   (2014-10-04)

YFP Coding Sequence

This construct contains the reporter protein coding sequence for the yellow fluorescent protein (YFP). It has been codon optimized for Staphylococcus aureus by Bose et al. (2013). YFP has an excitation peak at 514 nm and an emission peak at 527 nm (needs ref). We order the synthesis of this coding sequence from Biobasic. This is a reliable reporter in S.aureus (Malone et al., 2009).


YFP.png

The fluorescence and optical density of DH5α containing YFP was observed over time and compared to DH5α that did not contain YFP. Method: The expression cassette (in the pSB3K3 backbone) contained the YFP coding sequence under the constitutive sarA promoter BBa_K1323021 and TIR ribosome binding site BBa_K1323016. DH5α either containing the YFP plasmid or containing no plasmids were grown in triplicate overnight at 37°C in LB supplemented with the appropriate antibiotics. Cultures were diluted 1/100 into fresh LB with appropriate antibiotics and incubated at 37°C. Every hour, the fluorescence (excitation: 485 nm, emission: 535 nm) and optical density at 600 nm of 200 μl of culture was read using a plate reader.

Conclusion: The YFP coding sequence is able to produce a fluorescent protein.


Sequence and Features

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]


References

Bose, J. L., Fey, P. D., and Bayles, K. W. (2013). Genetic Tools to Enhance the Study of Gene Function and Regulation in Staphylococcus aureus. Applied and Environmental Microbiology. 79 (7): 2218-2224.


[edit]
Categories
//cds/reporter/yfp
//function/reporter/fluorescence
Parameters
colorYellow