Despite originally planning to work with X. laevis embryos this summer, my research took a new turn when the summer actually started. Now I am working towards a completely different goal: instead of looking at embryonic development and life, I’m looking at phages, which are viruses that can kill bacteria. Specifically, our lab is interested in the regulatory regions of these phages’ genomes, and understanding how that regulation can be strong or weak relative to its eventual output as proteins.
My research started out disappointing, with a tool we needed not working correctly. Our lab had received a plasmid (a circular loop of DNA that can enter a bacterium and be expressed there, provided the bacterium recognizes its regulation) from another lab. That plasmid was supposed to contain a reporter gene, a fluorescent marker which would glow if expressed. It was also promoterless, meaning that we could insert a regulatory region – in theory – and if the regulatory region was recognized by the host bacterium, the fluorescent protein would be expressed.
However, we ran into a lot of trouble trying to isolate this plasmid. There are essentially two ways to amplify a plasmid. We can use PCR, a technique which copies the plasmid outside of an organism, or we can grow up the plasmid inside many millions of bacteria and extract it from them.
We tried both methods. While the extraction of the plasmid from the host bacteria seemed initially successful, the story quickly fell apart, over a month of progressively more and more confusing data – which I will elaborate upon in my next post.