Dr. Amanda Hughes
The discussion started with us providing Dr. Hughes with an overview of our project, to which Dr Hughes gave her comments/suggestions. She remarked that to get an accurate idea of the UTR expression levels, we should measure the half-lives of the mRNA along with measuring the fluorescence protein expression. She even suggested we try and cut pieces of UTRs with higher half-lives and try and place different secondary structures at various points and measure their expression. Dr. Hughes also suggested that we should delve into conditional expression and include it in our project aspects and its potential application and establishing a proof of concept. She felt it was a very promising stream of thought to work upon and thought that we could even develop small molecules that are sort of DNA chaperones that could trigger conditional expression. The discussion continued, mentioning replicating the “guanine riboswitch” using our 5’ UTRs. She also talked about how we can develop systems that can change their half-lives based on either external conditions, temperature or by initiation sparked by small DNA chaperones. It was a very enlightening conversation, and it gave us new perspectives towards further building upon our work. Unfortunately, we couldn’t follow through with some of her suggestions, namely the measurement of mRNA half-lives on resource unavailability. However, we did follow through with her direction concerning using secondary structure scaffolds and were able to look at UTRs of heat shock and cold shock proteins. In fact based on her suggestions we set out to look for cold shock protein UTRs and found a very promising UTR called csPF the mutants of which we have taken forward after our learning stage to the second engineering cycle. We are highly grateful to Prof Amanda for her suggestions.
