Part:BBa_K526002
Temperature dependent Ribo-Switch fused to GFP
Previosuly, Olga et al (2011) described a temperature dependent riboswitch involved in regulating bacterial chemotaxis at high temperature (Fig 1A). They reported that the mRNA carrying the riboswitch was stable at all temperatures however, the protein expression was favored only at low temperature indicating a robustness in regulating bacterial chemotaxis (Fig 1B and Fig 1C).
We believe that this low-temperature dependent riboswitch would be of great interest in heterologous protein expression. Favoring expression only at a low temperature would gurantee that the heterologous proteins are folded properly without the formation of inclusion bodies.
We have exploited the temperature dependent riboswitch that is associated with the chemotaxis response regulator, CheR, to develop a novel temperature regulated gene expression system. The mRNA is assumed to form a stable secondary structure burrying the RBS and thus blocking gene expression at a higher temperature (37 degree C) where as the temperature is lowered to about 30 to 27 degree C the stability of the secondary structure is disturbed thus exposing the RBS and favoring gene expression.
In 2011 iGEM, we tried to engineer low temperature sensor into E. coli. As is shown in the above figure, when GFP was fused with a temperature dependent riboswitch (Fig 2B) it could sense low temperature: There was no GFP expression at 37 degree celsius. But a significant GFP expression was found at 30 degree celsius indicating the functionality of the ribo-switch.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 630
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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