Part:BBa_K613000
Medium-strength Plac promoter
This is a medium-strength Plac promoter. Placed before a gene, it promotes its expression. Plac is repressed by LacI. This promoter can be used for in vivo reporter systems; its medium strength is interesting for reporter genes that have a long half-life or that are not well supported by the cell.
Characterization
This promoter was characterized by coupling it to RFP; the Plac-RFP complex was inserted into the pSB3K1 plasmid and transformed into DH5aplha cells. Cells were grown in a medium containing Kanamycin plus different concentrations of IPTG, ranging from 0 to 1000 µM. OD600 absorbence and RFP fluorescence were measured every 10 minutes during 12 hours on a platereader machine.
IPTG induction
Fluorescence measurements (RFUs) were normalized by OD600 values.
The cells treated with IPTG produced more RFP than their non-treated counterparts; this difference is not striking, showing that the basal expression of LacI by our DH5alpha cells is quite low. The normal expression of RFP driven by our Plac promoter is of 530 normalized RFUs; the highest value is 640 normalized RFUs.
Dose-response curve
Fluorescence measurements (RFUs )were normalized by OD600 values. For each IPTG concentration, we estimated the steady-state fluorescence expression by averaging the measurements over the last hour.
This graph shows the action of IPTG on RFU output. Since it doesn't seem to saturate, there was perhaps still some repression by LacI occurring in our cells. Interestingly, LacI does have a quantifiable action on our Ptet promoter.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
| None |