Part:BBa_K199095:Experience

We tested K199095 (with the suppressor tRNA downstream of the RFP) using IPTG to induce the pLac promoter in this device. The positive control is Part:BBa_J04450. The negative controls are cells lacking RFP, and K199095 with no IPTG. When quantified, we found K199095 with IPTG to produce 4% as much RFP as J0445. A production of RFP indicates the suppressor tRNA carrying the anticodon of CCACU was able to suppress the 5mer frame shift mutation. It is worth noting that K199095 suppressed twice as much as its sister device of Part:BBa_K199094 which has the same tRNA and promoter but the RFP is downstream of the tRNA.

We think that having the tRNA downstream of the RFP simply allows two "doses" of promoter to increase the production of suppressing tRNA. This hypothesis is supported by the fact that when this same tRNA was used with pBad promoter (Part:BBa_K199058), the cells produced no detectable RFP. When the promoter was changed to the stronger pTet in part K199094, RFP was detected.

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