Difference between revisions of "Part:BBa K3084000"

 
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<partinfo>BBa_K3084000 short</partinfo>
 
<partinfo>BBa_K3084000 short</partinfo>
  
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Our team wanted to engineer the genome of an S-TIP37 phage to express a protein upon infection. In order to have the best expression possible, we looked into the proteins in the phage genome that are strongly expressed.  
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This Venus YFP gene has been designed for optimal translational efficiency from the genome of bacteriophage S-TIP37 during infection of its host, Synechococcus sp. strain WH8109. The translation of this gene is the same as the Venus YFP gene (NCBI accession number: ACQ43942). To maximise translational efficiency during S-TIP37 infection, the codon distribution of the S-TIP37 capsid gene, putatively the most strongly expressed gene on the S-TIP37 genome, was applied to the Venus YFP gene.
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Since translational efficiency is a strong determinator of the scale of expression, yellow fluorescent protein was codon-optimized for S-TIP37 phage capsid protein, in order to obtain a strong expression upon infection of Synecoccocus WH8109
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Comparing the translational efficiency to other YFPs in the S-TIP37 expression system required an experimental setup that was too complicated for the scope of our summer iGEM project. However, the fluorescence of our codon optimised gene was successfully tested using the pET expression system in E. Coli. Proof of fluorescence is demonstrated in figure 1.
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[[File:T--KU LEUVEN--Fluorescence Venus.jpg]]
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Optimal excitation wavelengths were determined by Sarkar et al. and found to be: 285 and 470 nm.
  
 
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===Usage and Biology===
 
===Usage and Biology===
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This part is useful as a reporter, most suitable for expression during Synechococcus infection by phage S-TIP37.
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===References===
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[https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2754229/] Sarkar, P., Koushik, S. V., Vogel, S. S., Gryczynski, I., & Gryczynski, Z. (2009). Photophysical properties of Cerulean and Venus fluorescent proteins. Journal of biomedical optics, 14(3), 034047. doi:10.1117/1.3156842
  
 
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Revision as of 19:06, 18 October 2019


Venus Yellow Fluorescent Protein (YFP), S-TIP37 codon optimised

This Venus YFP gene has been designed for optimal translational efficiency from the genome of bacteriophage S-TIP37 during infection of its host, Synechococcus sp. strain WH8109. The translation of this gene is the same as the Venus YFP gene (NCBI accession number: ACQ43942). To maximise translational efficiency during S-TIP37 infection, the codon distribution of the S-TIP37 capsid gene, putatively the most strongly expressed gene on the S-TIP37 genome, was applied to the Venus YFP gene.

Comparing the translational efficiency to other YFPs in the S-TIP37 expression system required an experimental setup that was too complicated for the scope of our summer iGEM project. However, the fluorescence of our codon optimised gene was successfully tested using the pET expression system in E. Coli. Proof of fluorescence is demonstrated in figure 1.

T--KU LEUVEN--Fluorescence Venus.jpg

Optimal excitation wavelengths were determined by Sarkar et al. and found to be: 285 and 470 nm.

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 424
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 682
  • 1000
    COMPATIBLE WITH RFC[1000]