Team:SJTU-BioX-Shanghai/Project/Subproject1/Results 1

Rare-Codon Switch

tRNA Modulator + Reporter for Qualitative Analysis

Results

We have designed PT7-RFP-6AGG ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K567017 BBa_K567017]) for qualitative analysis of tRNA modulator. The function of tRNA Modulator is tested by observing red fluorescence in the expression system.

In the experiment group, ER2566 is co-transformed with both Modulator lacI-Ptrc-tRNA^Arg ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K567001 BBa_K567001]) and Reporter PT7-RFP-6AGG ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K567017 BBa_K567017]). When Modulator is induced, tRNA^Arg is overexpressed, ribosome can get through the 6 consecutive AGG codons, RFP can be successfully expressed. For the control group in which only Reporter is transformed, RFP expresssion is hindered. The test results are as follows.

Fig 1. Confocal Microscope examining function of tRNA Modulator + Reporter for Qualitative Analysis.

Confocal Microscope analysis showed that RFP expression is hindered by the six consecutive AGG codons. After tRNA^Arg expresson is induced, RFP-6AGG expression is ensured.

Fig 2. tRNA Modulator + Reporter for Qualitative Analysis. tRNA Modulator + RFP-6AGG(the middle three) emit bright red fluorescence as wild type RFP(first one from the left). Control(first one from the right) exhibits no red fluoresence.

Conclusions

We have successfully constructed the tRNA Modulator and Reporter for qualitative analysis and tested their functions. Our results showed that through overexpression of rare tRNA, tRNA Modulator is indeed an excellent device for turning up protein biosynthesis for Reporter with rare codon insertions.