Team:Rice/Model

Motivation

Existing RNA thermometers operate around 37°C and do not exhibit significant conformational changes between 25°C and 30°C. As most plants will die at 37°C, it was necessary to design thermometers that experienced conformational change between 25°C. We chose to design thermometers by finding optimal candidates using a genetic algorithm, as the complexity of RNA folding made it difficult to rationally design thermometers. As the figures below show,A. Thaliana grows best between 18-20°C and P. putdia is typically grown in lab at 30°C. These two facts combine help strengthen the need to design RNA thermometers that are optimized to melt at 30°C.

How RNA Thermometer Regulate Translation

In essence, RNA thermometers are a form of temperature dependent translational regulation. At low temperatures, there is a higher probability of more base pairs forming what is known as a "stem-loop structure". At higher temperatures, the thermometers "melt", meaning there is a decreased likelihood of base pairs forming.

RNA Thermometer Design

The stem-loop structure of the thermometer was created by taking the complement of the ribosome binding site(RBS) and surrounding the nucleotides and introducing mutations into the sequence. The resulting altered sequence is known as the variable region. Mutating this region serves a two prong purpose. First off, it ensures that the RBS will be locked into the structure of the thermometer, rendering it inaccessible to the ribosome at low temperatures. Secondly, it allows us to optimize for the targeted melting temperature.